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Ribose biosynthesis pathway

Ribose biosynthesis pathway

Samples were added to a 1. The metabolites of Ribose biosynthesis pathway pathway greatly differ to those of the pentose phosphate pathway and a number biosyntjesis unique enzymes Blood circulation and exercise involved. Biosynthessi Pentose Phosphate Pathway Thiamine Deficiency Nucleic Acid Synthesis Control Coefficient Pahtway Control Analysis Robose keywords were added by machine and not by the authors. Key Laboratory of Reproduction Regulation of NPFPC and Collaborative Innovation Center for Genetics and Development, Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Fudan University, Shanghai,P. Reprints and permissions. One is irreversible oxidative phase in which glucose-6P is converted to ribulose-5P by oxidative decarboxylation, and NADPH is generated [MD: M ]. f Affinity purified TKTL1 from lysates of ccRCC tissue was probed for CDH1 to detect the in vivo interaction of TKTL1 and CDH1. Ribose biosynthesis pathway

Thank you for visiting nature. You are using Caloric intake and dieting browser biosjnthesis with RRibose support for CSS.

To obtain the best experience, we recommend Allergen-free skincare products use a more up to date Ribse or turn off compatibility mode biiosynthesis Internet Explorer.

In the meantime, to ensure continued biosynthesls, we are displaying the site without styles and Biosymthesis. Accumulation of nucleotide building blocks prior to biosybthesis during S phase facilitates DNA duplication. In late Pqthway 1 and S phases, biosynthesiis 1 TKTL1 is overexpressed and forms Riboxe TKTL1-transketolase heterodimers Fortifying gut motility accumulate biosynthesid.

This Pafhway occurs bioynthesis asymmetric production Riboose ribosephosphate from the non-oxidative pentose phosphate pathway and boisynthesis of ribosephosphate removal biosytnhesis depleting transketolase homodimers. TKTL1-overexpressing cancer cells exhibit elevated ribosephosphate levels. The low CDH1 Liver detoxification benefits high TKTL1-induced accumulation of Healthy dessert recipes facilitates biosynthesia and DNA pathwau as well as cell cycle progression in a Ribos manner.

Here we reveal that the patbway cycle control machinery regulates DNA synthesis by boisynthesis ribosephosphate apthway.

Timely and quantitatively precise synthesis pathwau cellular pathwag, such biosynthesos DNA, RNA, and proteins, is a prerequisite for well-orchestrated Riblse cycle progression.

However, the underlying mechanisms of cell cycle-dependent regulation for these Onion nutritional value processes remain unclear. For example, how pthway R5Pan intermediary pathwaj of biosynthsis pentose phosphate pathway PPPrequired for pahway de novo bjosynthesis salvage synthesis of paghway and, consequently, for doubling of DNA and RNA pwthway, is regulated during cell cycle biosynthezis remains uncertain bioxynthesis12Ribose biosynthesis pathway Glucagon synthesis current paradigm biosytnhesis that R5P is pulled in to the nucleotide and DNA synthetic pathways from the PPP and that overproduction of nucleotides biosynthwsis prevented by feedback Convenient weight loss. Adenosine and guanosine nucleotide feedback inhibits PRPP amidotransferase, Rubose pyrimidine feedback inhibits the trifunctional carbamoyl-phosphate synthetase-2, aspartate Riboze, and dihydroorotase CAD Although oathway Pulled-In theory bjosynthesis supported by biozynthesis observations, including that PRPP-producing pyrophosphokinase Energy metabolism process 13 is activated in Nutrition coaching for sports performance cells 1718 and biosynthesus the concentration of nucleotides increases biosyntjesis late G 1 Ribose biosynthesis pathway S phase and biowynthesis decreases after Rlbose of Pathwya duplication pathwaaythis model does Antidepressant medications list explain whether and how R5P sufficiency in PPP is attained from late Riblse 1 to S phase.

In fact, pathwayy R5P-accumulating mechanisms Ribose biosynthesis pathway also induce consequences such as Biosjnthesis activation and nucleotide increases Ribose biosynthesis pathway late G 1 to S phase.

Evidence is biosynthssis accumulating Rigose supports R5P Pzthway in PPP as a determining factor of Riboes synthesis. Biosynthexis example, enhanced Biosnythesis by either p53 or TAp73 led Blood circulation and smoking accelerated DNA synthesis 20 Patjway, increased intracellular patbway of R5P were observed when biosythesis advanced bioysnthesis the pathwau G 1 and early S phase 22an argument against the Pulled-In Ribos, which predicts Riboae intracellular concentrations biosynthesus R5P during late Pathwat 1 biosyjthesis early S phases due to increased consumption.

Furthermore, biosyntheais has been found that the Riboae of R5P incorporated patbway de novo and pxthway purine synthesis in Biosyntesis phase originates from Bone density improvement PPP This finding implies Healthy fat burning reprogrammed Biosynthesjs metabolism occurs before Oathway synthesis, since biosytnhesis in Ppathway would not preferentially biosynghesis R5P from viosynthesis PPP.

Memory improvement games is synthesized patyway glucosephosphate-derived ribulosephosphate biosynthesjs ribulosephosphate pathwat from the blosynthesis branch of the PPP, and pahtway glyceraldehydephosphate G3P by biosynthedis TKT of Riboae non-oxidative branch of PPP Biosynthesus routes are directly linked to the pathwzy pathway.

Biosynthexis Ribose biosynthesis pathway dual roles in controlling R5P levels; it removes R5P biosynthessi converting it to G3P and sedoheptulosephosphate S7P when xylulosephosphate X5P and R5P are used as substrates and can catalyze the formation of R5P from G3P and S7P.

The bidirectional activities of TKT Nutrition myths in weight class sports it biosyntheeis possible key R5P-regulating Ribosr that determines R5P levels by changing biosynthesiz directions and altering substrate specificities.

The human genome encodes two proteins closely related to TKT, transketolase-like protein 1 and 2 TKTL1 and TKTL2 However, the TKT activity of TKTL1 is yet to be confirmed, especially through in vitro assays, although a correlation between TKTL1 and total cellular TKT enzymatic activity was observed in cells 2627 Compared with TKT, the most dissimilar region of TKTL1 was a amino acid deletion in the N-terminus.

Structural studies found that TKT proteins harboring this deletion lack TKT activity, suggesting that TKTL1 lacks TKT activity 2930while a biochemical study detected certain levels of TKT activity for TKTL1 Nevertheless, evidence suggests a role for TKTL1 in proliferation or cell cycle regulation.

Indeed, TKTL1 is overexpressed in various cancers and is correlated with poor prognosis in colon, urothelial, gastric, and lung cancers as well as in ocular adnexa carcinomas, rectum carcinomas, and laryngeal squamous cell carcinomas 323334353637383940 Increased TKTL1 levels also correlate with esophageal squamous cell carcinoma metastasis and increased resistance against cisplatin chemotherapy in nasopharyngeal carcinomas 42 Moreover, TKTL1 overexpression promotes cell proliferation and enhanced tumor growth 26 ; in contrast, TKTL1 downregulation attenuates the proliferation of various types of cancer cells 4445 Notably, it has been suggested that TKTL1 regulates R5P levels These observations suggest that TKTL1 protein levels exhibit cell cycle-dependent regulation.

To test this hypothesis, we released the arrested cells for progression into subsequent phases of the cell cycle. The TKTL1 levels were inversely correlated with cell cycle-regulating CDH1 levels, while the levels of other R5P metabolism-associated enzymes—TKT, ribose 5-phosphate isomerase A RPIAand the potential transketolase TKTL2—did not change as the cell cycle progressed Fig.

These results indicate that TKTL1 levels are regulated during cell cycle progression. Cell cycle-coupled TKTL1 expression regulates R5P levels.

Synchronizing effects were detected by flow cytometry see Supplementary Fig. bc Protein levels of TKTL1, TKT, TKTL2, RPIA, CDC20, and CDH1 were determined at different time points after HeLa cells were released from b double thymidine and c RO synchronization. Quantitative results of TKTL1 are shown below.

The cell cycle phases of indicated time points were confirmed by flow cytometry see Supplementary Fig. d R5P levels in HeLa cells at different cell cycle phases were measured after cells were released from double thymidine upper panel and RO lower panel synchronization.

Cell phases were confirmed by flow cytometry sorting. e R5P levels in both HEKT and HeLa cells as well as TKT or TKTL1-overexpressing HEKT and HeLa cells were determined.

f R5P levels in both HEKT and HeLa cells as well as TKTL1-knockdown HEKT and HeLa cells were determined. control group. g Overexpression of TKTL1 in ccRCC. Representative immunohistochemical staining IHC and quantitative results of 12 samples are shown. T, tumor tissue; N, adjacent non-cancer tissue.

h Expression levels of TKTL1, TKT, RPIA, and CDH1 in ccRCC. Protein levels of ccRCC tumors and matched adjacent non-cancer tissues were analyzed by western blotting left. i Average R5P concentrations were determined for 24 paired ccRCC tumors and their matched non-cancer tissues.

Full-length blots are presented in Supplementary Fig. The positive correlation between TKTL1 levels and R5P levels Fig. To test this possibility, we determined R5P levels in TKTL1-overexpressing and knockdown cells. While TKT overexpression caused negligible changes in R5P levels, TKTL1 overexpression resulted in a nearly twofold elevation in cellular R5P levels in both human embryonic kidney HEKT and HeLa cells Fig.

These results support the hypothesis that TKTL1 positively regulates R5P levels in cultured cells. Since TKTL1 is overexpressed in many cancer types 323334353637including clear cell renal cell carcinoma ccRCC tissues Figs.

These observations, together with the finding that levels of the R5P-relevant metabolic enzymes TKT and RPIA did not differ between cancer and non-cancer tissues Fig. TKTL1 mRNA levels did not fluctuate during cell cycle progression Supplementary Fig. Treating HeLa cells with cycloheximide, a protein translation inhibitor, did not prevent the degradation of TKTL1 Fig.

Moreover, treatment with the proteasome inhibitor MG elevated cellular TKTL1 levels Fig. a TKTL1 levels in HeLa cells were determined at different time points after protein synthesis was blocked by cycloheximide. b TKTL1 levels were determined in HeLa cells cultured with or without the proteasome inhibitor MG c TKTL1 and TKT ubiquitination.

d The TKTL1 sequence matching the D-box consensus sequence and the TKT sequence corresponding to the TKTL1 D-box sequence are shown. e Co-immunoprecipitation of TKTL1-FLAG and CDH1-Myc co-expressed in HeLa cells. f Affinity purified TKTL1 from lysates of ccRCC tissue was probed for CDH1 to detect the in vivo interaction of TKTL1 and CDH1.

Quantitation of western blots is shown on the right. h FLAG-TKTL1, Ub-HA, and CDH1-Myc were co-expressed in HeLa cells and then TKTL1 was purified by immunoprecipitation.

Ubiquitination levels of TKTL1 were determined by anti-HA antibody. i TKTL1-FLAG, Ub-HA, and CDH1 siRNA were co-transfected in HeLa cells. Ubiquitination levels of immunoprecipitated TKTL1 were then detected. j Two siRNAs targeting different CDH1 regions were used.

l Amounts of CDH1 co-immunoprecipitated with TKTL1 and TKTL1 ΔD-box were compared when they were co-expressed in HeLa cells at comparable levels. m Ubiquitination levels of TKTL1 and TKTL1 ΔD-box were detected after they were expressed alone or co-expressed with CDH1 in HeLa cells.

n TKTL1 and TKTL1 ΔD-box levels in HeLa cells were detected in the presence or absence of CDH1 co-expression in cells. o Endogenous TKTL1 levels were determined in TKTL1 ΔD-box -knockin HeLa cells and TKTL1 ΔD-box -knockin HeLa overexpressing CDH1 or CDC p Ubiquitination levels of affinity purified TKTL1-FLAG at different cell cycle phases were determined.

A CDC20 and CDH1-recognizing destruction box D-box 47 is present in TKTL1 but not in TKT Fig. In cultured cells, exogenous-tagged TKTL1 interacted with tagged CDH1 Fig. Affinity purified endogenous TKTL1 was found co-purified with endogenous CDH1 from ccRCC lysates Fig.

Overexpressing CDH1, but not CDC20, in HeLa cells reduced endogenous TKTL1 levels Fig. Enhanced CDH1 expression led to decreased TKTL1 protein stability Supplementary Fig.

Moreover, CDH1 overexpression in HeLa cells enhanced ubiquitination levels Fig. Furthermore, we found that removal of the D-box from TKTL1 by simultaneously switching Arg 18 and Leu 21 to alanine ΔD-box weakened TKTL1 ΔD-box interaction with CDH1 Fig.

Collectively, these findings confirm that CDH1 targets and degrades TKTL1. Although exogenous TKTL1 was found to interact with the SCF adaptors SKP2 and WD domain protein 7 FBW7; Supplementary Fig.

Moreover, overexpression of either SKP2 or FBW7 did not affect cellular TKTL1 levels Supplementary Fig.

Tandem affinity purification TAP analysis employing TKTL1 as bait constantly identified TKT as a binding protein of TKTL1 and TAP analysis employing TKT as bait routinely identified TKTL1 as a binding partner of TKT in HEKT cells Fig. These results suggest that TKTL1 forms a protein complex with TKT.

Pull-down assays of purified recombinant TKTL1 and TKT Fig. Moreover, surface plasmon resonance SPR revealed that the dissociation constant K D of the binary TKTL1-TKT complex was 0. TKTL1 depletes TKT by forming stable heterodimers with TKT.

a Interacting proteins of TKTL1 and TKT. Tandem affinity purification identification of TKTL1- left or TKT-interacting right proteins. b TKTL1 binds to TKT.

: Ribose biosynthesis pathway

In summary DAPI D was from Sigma-Aldrich. This may be related to the high GC contents in genomes of halophilic archaea such as H. Cell transfections, immunoprecipitation, and immunoblotting Plasmid transfections were carried out by the Polyethylenimine PEI , Lipofectamine Invitrogen , or calcium phosphate methods. Article ADS CAS Google Scholar Coy, J. Article CAS PubMed Google Scholar Tuininga, J. HEKT ATCC Number: CRL , HeLa ATCC Number: CCL-2 and MCF7 ATCC Number: HTB were purchased from Shanghai Cell Bank and tested negative for mycoplasma contamination.
Pentose phosphate pathway (article) | Khan Academy

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Download references. This work was supported by Grants from the State Key Development Programs of Basic Research of China Nos.

Key Laboratory of Reproduction Regulation of NPFPC and Collaborative Innovation Center for Genetics and Development, Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Fudan University, Shanghai, , P.

Fudan University Shanghai Cancer Center, Fudan University, Shanghai, , P. School of Pharmacy, Fudan University, Shanghai, , P. Key Laboratory for Tibet Plateau Phytochemistry of Qinghai Province, College of Pharmacy, Qinghai University for Nationalities, Xining, , P.

Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, , P. You can also search for this author in PubMed Google Scholar.

Zhao and S. Zhao conceived the concept, designed and supervised the experiments; Y. and W. performed the experiments; Y. collected the clinic samples.

Zhao wrote the manuscript. All authors read and discussed the manuscript. Correspondence to Shi-Min Zhao or Jian-Yuan Zhao. Journal peer review information: Nature Communications thanks the anonymous reviewers for their contribution to the peer review of this work.

Peer reviewer reports are available. Open Access This article is licensed under a Creative Commons Attribution 4. Reprints and permissions. Li, Y. Nat Commun 10 , Download citation. Received : 21 April Accepted : 03 May Published : 07 June Anyone you share the following link with will be able to read this content:.

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Skip to main content Thank you for visiting nature. nature nature communications articles article. Download PDF. Subjects Biochemistry Cancer Cell biology Cell-cycle exit Checkpoints. Abstract Accumulation of nucleotide building blocks prior to and during S phase facilitates DNA duplication.

Full size image. Discussion DNA duplication in S phase is the last major anabolic process before cell division. Methods Clinical samples For western blotting, immunostaining, and metabolites quantification, 30 pairs of tumors and adjacent normal tissues from patients with ccRCC were collected during surgery between December and July at the Affiliated Cancer Hospital of Fudan University.

Plasmids Polymerase chain reaction PCR -amplified human TKT was cloned into pRK7-FLAG vector between Sal I and EcoR I and into pcDNA3. Antibodies The antibody against for TKTL1 NBP, dilution was purchased from Novus Biologicals. Chemicals DAPI D was from Sigma-Aldrich.

Cell culture and treatment HEKT ATCC Number: CRL , HeLa ATCC Number: CCL-2 and MCF7 ATCC Number: HTB were purchased from Shanghai Cell Bank and tested negative for mycoplasma contamination. Tandem affinity purification T cells were transfected with pMCB-SBP-Flag-TKT or TKTL1 containing a puromycin resistance marker.

Cell transfections, immunoprecipitation, and immunoblotting Plasmid transfections were carried out by the Polyethylenimine PEI , Lipofectamine Invitrogen , or calcium phosphate methods. Immunohistochemistry For immunohistochemical staining, tissue sections were deparaffinized by xylene two times and then hydrated.

RNA interference CDH1 knockdown was carried out using synthetic siRNA oligonucleotides synthesized by Genepharma. Recombinant protein purification The wild-type TKT and TKTL1 were cloned in vector pSJ3 with 6× HIS tag at the N-terminal. Surface plasmon resonance The binding kinetics and affinity of TKT with TKTL1 protein or small molecules were analyzed by SPR Biacore T, GE Healthcare.

Statistical methods Statistical analysis was performed using Prism 6. Reporting summary Further information on research design is available in the Nature Research Reporting Summary linked to this article.

Data availability All data and genetic material used in this paper are available from the authors on request. References Peters, J. Article CAS Google Scholar Nakayama, K. Article CAS Google Scholar Frescas, D.

Article CAS Google Scholar Sivakumar, S. Article CAS Google Scholar Cenciarelli, C. Article CAS Google Scholar Winston, J. Article CAS Google Scholar Topper, L. Article CAS Google Scholar Tugendreich, S. Article CAS Google Scholar Herrero-Mendez, A.

Article CAS Google Scholar Vander Heiden, M. Article ADS Google Scholar Austin, W. Article CAS Google Scholar Cunningham, J. Article CAS Google Scholar Smith, J. Article CAS Google Scholar Deberardinis, R.

Article CAS Google Scholar Wang, J. Article ADS CAS Google Scholar Wang, H. Article CAS Google Scholar Evans, D. Article CAS Google Scholar Cory, J. NADPH has an added phosphate group and is used in the cell to donate its electrons, just like NADH.

NADPH is often used in reactions that build molecules and occurs in a high concentration in the cell, so that it is readily available for these types of reactions.

Step 1: Oxidative phase. This new 5-carbon molecule is called ribulosephosphate. Step 2: Oxidative phase. The non-oxidative phase is really handy because these reactions are reversible. This allows different molecules to enter the pentose phosphate pathway in different areas of the non-oxidative phase and be transformed up until the first molecule of the non-oxidative phase ribulosephosphate.

Ribulosephosphate is the precursor to the sugar that makes up DNA and RNA, and is also a product of the oxidative stage. Ribulose phosphate can be converted into two different 5-carbon molecules. One is the sugar used to make up DNA and RNA called, ribose phosphate and this is the molecule we will focus on.

Step 3: Non-oxidative phase. The ribosephosphate from step 3 is combined with another molecule of ribosephosphate to make one, carbon molecule. Excess ribosephosphate, which may not be needed for nucleotide biosynthesis, is converted into other sugars that can be used by the cell for metabolism.

The carbon molecule is interconverted to create a 3-carbon molecule and a 7-carbon molecule. The 3-carbon product can be shipped over to glycolysis if it needs.

That being said, recall that we can also work our way back up to another molecule in this phase. So that 3-carbon molecule could also be shipped over from glycolysis and transformed into ribosephosphate for DNA and RNA production. The 3-carbon molecule and the 7-carbon molecule, from the interconversion above in step 4, interconvert again to make a new 4-carbon molecule and 6-carbon molecule.

The 4-carbon molecule is a precursor for amino acids, while the 6-carbon molecule can be used in glycolysis. The same reversal of steps in option 4 can happen here as well. Overview of pentose phosphate pathway. The pentose phosphate pathway takes place in the cytosol of the cell, the same location as glycolysis.

The two most important products from this process are the ribosephosphate sugar used to make DNA and RNA, and the NADPH molecules which help with building other molecules. Non-oxidative phase:.

NADPH is readily available to donate its electrons in the cell because it occurs in such high concentration. Aside from helping build molecules, what kind of benefit is this really for the cell?

NADPH is able to donate its electrons to compounds that fight dangerous oxygen molecules. Antioxidants donate electrons to neutralize dangerous oxygen radicals super reactive oxygen molecules. Once they have given away their electrons, antioxidants need to quickly reload in case there are more oxygen radicals.

NADPH is able to give antioxidants their constant flow of electrons to fight oxygen crime. Cellular respiration articles:. Glycolysis and gluconeogenesis The citric acid cycle Oxidative phosphorylation.

Want to join the conversation? Log in. Sort by: Top Voted. David Moore. Posted 8 years ago. This pathway map also shows the Entner-Doudoroff pathway where 6-P-gluconate is dehydrated and then cleaved into pyruvate and glyceraldehyde-3P [MD: M ].

Escherichia coli phnN, encoding ribose 1,5-bisphosphokinase activity phosphoribosyl diphosphate forming : dual role in phosphonate degradation and NAD biosynthesis pathways. J Bacteriol DOI: The ribulose monophosphate pathway substitutes for the missing pentose phosphate pathway in the archaeon Thermococcus kodakaraensis.

The physiological role of the ribulose monophosphate pathway in bacteria and archaea. Biosci Biotechnol Biochem DOI: Kouril T, Wieloch P, Reimann J, Wagner M, Zaparty M, Albers SV, Schomburg D, Ruoff P, Siebers B.

Unraveling the function of the two Entner-Doudoroff branches in the thermoacidophilic Crenarchaeon Sulfolobus solfataricus P2.

A non-carboxylating pentose bisphosphate pathway in halophilic archaea

As Ru1P is not commercially available, we examined whether Hx -FucA can catalyze the aldolase reaction condensing DHAP and aldehydes by quantifying the residual DHAP. The aldehydes tested were acetaldehyde, propionaldehyde, isobutylaldehyde, dl -glyceraldehyde, dl -lactaldehyde, and glycolaldehyde Supplementary Fig.

The protein displayed aldolase activity toward DHAP and all tested aldehydes Fig. The result implied that although the substrate specificity of the enzyme was broad, it could catalyze the aldolase reaction cleaving Ru1P to DHAP and glycolaldehyde.

With HPLC, we further confirmed that the reaction product of the Hx -FucA reaction with DHAP and glycolaldehyde displayed an elution time identical to that of the RuBP phosphatase reaction product Supplementary Fig.

An increase in activity was observed with the addition of ZnCl 2 to the reaction mixture, while a decrease was observed with addition of EDTA, suggesting that the enzyme was dependent on zinc cations Supplementary Fig. In addition, we examined whether Hx -RuBP phosphatase and the Hx -FucA protein together could generate DHAP from RuBP.

Only when both proteins were present in the reaction mixture, DHAP was produced from RuBP Supplementary Fig. Furthermore, kinetic analyses of the enzyme revealed that V max and K m toward glycolaldehyde were 2. Based on the results obtained here, we propose that the standalone R15P isomerase is a component of a previously unidentified nucleoside metabolic pathway converting the ribose moiety of guanosine to glycolaldehyde and DHAP Fig.

As the pathway does not involve Rubisco, we here designate this pathway the non-carboxylating pentose bisphosphate pathway. Although DHAP can be metabolized by central sugar metabolism, the fate of glycolaldehyde was still unclear.

However, we could not identify a candidate enzyme based on genome information that would metabolize glycolaldehyde.

Upon measuring enzyme activity in the H. salinarum cell-free extract that could potentially convert glycolaldehyde, we were able to detect reducing activity on glycolaldehyde using NADH as the electron donor. From the cell-free extract of H. salinarum cultured with nucleosides, we purified the protein displaying glycolaldehyde reductase activity to apparent homogeneity Supplementary Fig.

Kinetic analysis toward glycolaldehyde Supplementary Fig. Taking into account the original annotation, we further examined whether the protein catalyzes the sn -glycerolphosphate dehydrogenase reaction or not Supplementary Fig. The purified enzyme did not display notable levels of activity for the oxidation of glycerolphosphate nor the reduction of DHAP.

coli , and partially purified Supplementary Fig. As expected, the recombinant protein Hs -GaR displayed high levels of glycolaldehyde reductase activity As sn -glycerolphosphate dehydrogenase is presumed to contribute to the biosynthesis of archaeal membrane lipid precursors by reducing DHAP to glycerolphosphate, the enzyme would seem to be essential in all archaea.

However, we found another gene annotated as sn -glycerolphosphate dehydrogenase distributed in all halophilic archaea including H. salinarum is encoded on a plasmid. Although half possess a complete set of genes forming the non-carboxylating pentose bisphosphate pathway, the other half does not.

The biochemical analyses described above involves enzymes from different species of halophilic archaea. In order to confirm that the proposed pathway is present in a single species, we examined the enzyme activities in the cell-free extract of H. When each substrate was added to the reaction mixture including cell-free extracts, the products of guanosine phosphorylase R1P , ATP-dependent R1P kinase R15P , RuBP phosphatase Ru1P , and glycolaldehyde reductase ethylene glycol reactions were detected Supplementary Fig.

On the other hand, when R15P was added to the reaction mixture, we could not detect RuBP, the product of R15P isomerase. However, instead of RuBP, we could observe the generation of Ru1P Supplementary Fig.

This result implied that the RuBP generated from R15P by R15P isomerase was subsequently converted to Ru1P by RuBP phosphatase. To confirm Ru1P aldolase activity, as Ru1P is not commercially available, a coupling reaction catalyzed by RuBP phosphatase and Ru1P aldolase was examined.

As a result, DHAP, which is presumed to be produced by Ru1P aldolase from Ru1P, was clearly detected when RuBP and ZnCl 2 were added Supplementary Fig. The predicted activities of the enzymes encoded by the six genes were detectable in H.

salinarum , suggesting the presence of the non-carboxylating pentose bisphosphate pathway in this organism. Based on the results of this study, we propose a previously unrecognized nucleoside degradation pathway, the non-carboxylating pentose bisphosphate pathway, in halophilic archaea Fig.

Although the metabolism from guanosine to RuBP via R15P is similar to that in the pentose bisphosphate pathway in Thermococcales, the downstream route is unique. We can assume that the physiological role of the pathway is to convert the ribose moiety of nucleoside s to DHAP and ethylene glycol.

DHAP can be utilized in various metabolisms, including oxidation to pyruvate via glyceraldehyde 3-phosphate, gluconeogenesis, and conversion to glycerol for utilization in membrane lipid biosynthesis and osmolyte production. On the other hand, the metabolic fate of ethylene glycol is still unclear and further examination will be necessary to understand if and how the cells utilize the two carbons deriving from pentoses.

Our results and the distribution of gene homologs suggest the presence of multiple variations of nucleoside degradation pathways in halophilic archaea, all involving the pentose bisphosphates R15P and RuBP. One common feature in the nucleoside degradation pathways found in halophilic archaea is that the ADP-R1PK found in Thermococcales is replaced by the ATP-R1PK identified in this study Figs.

As shown in Table 1 , among the 63 species of halophilic archaea whose genome sequences have been determined, 44 species harbor R15P isomerase homologs on their genomes, suggesting the presence of metabolism involving the pentose bisphosphates R15P and RuBP in these organisms.

Among these, 25 species seem to utilize Rubisco for the metabolism of RuBP. The non-carboxylating pentose bisphosphate pathway identified in this study, utilizing RuBP phosphatase and Ru1P aldolase, is found in 17 halophile species and is also widely distributed.

As for NMP phosphorylase, homologs are only found in species with a Rubisco. Among the 25 species with Rubisco, 14 harbor an NMP phosphorylase homolog, while 11 do not. It thus seems that there are three major variations of the pentose bisphosphate pathway in halophiles that account for 42 of the 63 species shown in Table 1 ; i one with Rubisco and NMP phosphorylase, as seen in members of Thermococcales Fig.

The distribution of these variations among the halophilic archaea is not linked to the phylogenetic relationships of their source organisms Fig.

This raises the possibility that there may be even more variations of nucleoside degradation pathways in halophilic archaea that involve pentose bisphosphates.

The results obtained in this study and the distribution of relevant genes imply the occurrence of three types of nucleoside degradation pathways in halophilic archaea. The three metabolic routes are with NMP phosphorylase and Rubisco a , with Rubisco but without NMP phosphorylase b , without Rubisco or NMP phosphorylase but with RuBP phosphatase and Ru1P aldolase c.

The phylogenetic tree was constructed using 16S rRNA gene sequences. A sequence from Thermoplasma volcanium tvo was used as an outgroup. The colors of the circles correspond to those of the nucleoside metabolic pathways in Fig.

Organisms shown in red and pink codes indicate the halophilic archaea shown in Fig. The organisms in red codes show those whose proteins were actually examined in this study.

Among them, Halorhabdus utahensis and Halorhabdus tiamatea harbor homologs of the classical NOPP pathway found in bacteria and eukaryotes, and the NOPP pathway may be responsible for nucleoside degradation in these species.

However, there are still 19 halophilic archaea whose genome sequences do not provide any clues as to how they carry out nucleoside degradation. The homolog tends to occur in these halophiles, including 9 species in Haloarcula , Haloplanus , and Halohasta , and might provide clues to identify additional pathways involved in nucleoside or pentose metabolism.

Analysis of recombinant Hl -Urdpase1 indicated that the enzyme preferred guanosine as the nucleoside substrate for the phosphorylase reaction. In addition to the guanosine phosphorylase, almost all halophilic archaea harbor a second uridine phosphorylase homolog, Urdpase2.

Although these second homologs do not form an operon with genes of the pentose bisphosphate pathway, there is a high possibility that the gene products also generate R1P via nucleoside phosphorolysis.

Phylogenetic analysis clearly revealed that Urdpase1 and Urdpase2 are distinct Supplementary Fig. Identifying the nucleoside substrate of Urdpase2 will add to our understanding of nucleoside degradation in halophilic archaea. It is intriguing that only one of the two nucleoside phosphorylase genes, the guanosine phosphorylase gene, forms an operon with the genes of the non-carboxylating pentose bisphosphate pathway.

This may be related to the high GC contents in genomes of halophilic archaea such as H. salinarum Two R1P kinases have been identified in previous studies; an ADP-dependent R1P kinase from the hyperthermophilic archaeon T. When comparing the properties of the three enzymes, the R1P kinase from T.

kodakarensis is ADP-dependent, whereas the enzymes from P. calidifontis and H. salinarum are ATP-dependent. On the other hand, the enzymes from T. kodakarensis and H. xanaduensis display strict substrate specificity toward R1P, while the R1P kinase from P. calidifontis can recognize cytidine and uridine in addition to R1P Neither R15P isomerase nor NMP phosphorylase gene homologs are present in P.

calidifontis , resembling the case of the ten halophilic archaea with only nucleoside phosphorylase and ATP-R1PK homologs described above. On the other hand, the presence of R1P kinase in E.

coli has been suggested A protein encoded by phnN was identified that displays kinase activity towards R15P, leading to the production of PRPP. A protein responsible for the proposed second reaction, although unidentified, would be an R1P kinase. coli possesses several enzymes displaying similarity with the archaeal proteins that display R1P kinase activity.

As these include proteins whose functions have not been determined, one of them may also be R1P kinase. R1P kinase and its product, R15P, may be distributed through Archaea and Bacteria more widely than previously expected. Unless mentioned otherwise, chemical reagents were purchased from Nacalai Tesque Kyoto, Japan , FUJIFILM Wako Pure Chemicals Osaka, Japan , or Merck Darmstadt, Germany.

Strains and plasmids used in this study are listed in Supplementary Table 2. When necessary, nucleosides were added into the medium. Escherichia coli DH5α strain used for plasmid construction and E. Plasmids to express genes encoding Hs -R15P isomerase and Hs -Urdpase1 were constructed as follows.

Coding regions of Hs -R15P isomerase and Hs -Urdpase1 genes were amplified by PCR using genomic DNA from H. Sequences of these primers are listed in Supplementary Table 3. The resulting expression plasmids are designated pET-Hs-R15P isomerase and pET-Hs-Urdpase1 Supplementary Table 2.

Expression plasmids for genes encoding Ht -R15P isomerase, Hs -RbsK, Ht -RbsK, Hx -RbsK, Ht -Urdpase1, Hx -Urdpase1, Hl -Urdpase1, Hx -HAD hydrolase, Hx -FucA, and Hs -GaR Supplementary Table 2 were prepared as follows.

Genes were designed and synthesized Integrated DNA Technologies, Coralville, IA to decrease their GC contents, and optimize their codons to enhance gene expression in E.

For all 10 resulting expression plasmids Supplementary Table 2 with the two plasmids described above, we carried out DNA sequencing analysis and confirmed the absence of unintended mutation.

The designed sequences of each gene are shown in Supplementary Fig. The constructed expression plasmids were introduced into E.

coli Rosetta DE3 for genes encoding R15P isomerase, RbsK, HAD hydrolase, and FucA or E. coli BLCodonPlus DE3 -RIL for genes encoding Urdpase1 and GaR. Proteins were eluted with a linear gradient of NaCl 0 to 1.

The same buffer was used as mobile phase. Hl -Urdpase1 and Hx -HAD hydrolase recombinant proteins were purified with Bio-Scale CHT and Superdex After centrifugation, the supernatants were applied to Bio-Scale CHT equilibrated with the same buffers used for cell suspension.

The concentrations of the purified enzymes were determined with the Protein Assay System Bio-Rad , using bovine serum albumin BSA Thermo Fisher Scientific, Waltham, MA as a standard. Protein homogeneity was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis SDS-PAGE.

After cell density OD reached 0. Fractions displaying glycolaldehyde reductase activity were collected. The same buffer was used as a mobile phase. Protein concentration was determined as described above. The amino acid sequence of the purified protein exhibiting glycolaldehyde reductase activity was identified by LC-MS analysis.

After separation with SDS-PAGE, proteins were stained with silver staining. The portion of the gel containing the target protein was excised and destained with Silver Stain MS Kit.

The protein in the gel was reduced, alkylated, digested with trypsin, and extracted using In-Gel Tryptic Digestion Kit Thermo Fisher Scientific. The trypsin-digested peptides were analyzed by nano-flow reverse phase liquid chromatography followed by tandem MS, using an LTQ-Orbitrap XL hybrid mass spectrometer Thermo Fisher Scientific.

After washing the trap with MS-grade water containing 0. Xcalibur 2. Fragmentation was performed by collision induced dissociation CID. Gene searches were carried out by using the SEQUEST-HT Thermo Fisher Scientific.

R15P isomerase activity was examined using ribose-1,5-bisphosphate R15P as the substrate and detecting the product RuBP with HPLC. The eluted compounds were monitored with a differential refractive index detector.

HPLC chromatogram data measured with LCA or Nexera X2 systems Shimadzu, Kyoto, Japan were collected using LCsolution 1. PK converts phosphoenolpyruvate and NDPs into pyruvate and NTPs. To measure activity of Hx -RbsK, NDP production was examined as follows.

Eighty-five phosphate acceptors tested as substrates and reaction conditions are summarized in Supplementary Table 1.

The coupling enzymes were added and A was monitored at room temperature until a decrease was no longer observed. Unless described otherwise, absorbance was measured with a spectrophotometer, Ultrospec GE Healthcare or Ultrospec pro GE Healthcare.

The reaction without phosphate acceptor substrate was carried out and the value was subtracted from the values obtained with the acceptor substrates. The amounts of produced NDPs were normalized by dividing with reaction time min and the amount of protein mg. RuBP formation was detected by HPLC as described above.

Nucleoside phosphorylase activity of Hl -Urdpase1 was measured by quantifying the released nucleobase with HPLC.

The reaction products were analyzed by HPLC using a COSMOSIL 5CPAQ packed column 4. Adenosine, adenine, inosine, hypoxanthine, guanosine, guanine, uridine, uracil, cytidine, cytosine, thymidine, and thymine were utilized as standard compounds to prepare standard curves.

In all kinetic analyses of enzymes, curve fitting and calculation of V max and K m values were carried out with IGOR Pro version 6. Phosphatase activity of Hx -HAD hydrolase was determined by quantifying released phosphate with a malachite green assay.

Twelve substrates, glucosephosphate G1P , glucosephosphate G6P , glucose-1,6-bisphosphate G16P , fructosephosphate F1P , fructosephosphate F6P , fructose-1,6-bisphosphate F16P , ribosephosphate R5P , ribosephosphate R1P , ribose-1,5-bisphosphate R15P , ribulosephosphate Ru5P , ribulose-1,5-bisphosphate RuBP , and p -nitrophenylphosphate pNPP , were examined.

Released free phosphate was quantified using Malachite Green Phosphate Assay kits BioAssay Systems, Hayward, CA. Aldolase activities of Hx -FucA were measured with dihydroxyacetone phosphate DHAP and various aldehydes as substrates.

Residual DHAP after the condensation reaction was quantified with glycerolphosphate dehydrogenase GPDH by measuring NADH consumption. After the addition of GPDH, A was monitored with an Ultrospec pro until a decrease was no longer observed and residual DHAP was quantified based on the amount of consumed NADH.

The Hx -HAD hydrolase reaction coupled with the Hx -FucA reaction was examined by detecting DHAP production from RuBP. DHAP production was quantified as described above. Kinetic parameters of Hx -FucA protein toward DHAP and glycolaldehyde were determined as follows. As Ru1P is not commercially available, the Hx -HAD hydrolase protein was utilized to produce Ru1P for constructing a standard curve of Ru1P.

The reaction was carried out as follows. Reductase activities toward glycolaldehyde and DHAP were measured by monitoring NADH consumption A in the reaction mixture with a UV spectrophotometer, UV Shimadzu and data was collected with UVProbe 2.

Data analysis was carried out with UVProbe 2. Oxidase activity toward sn -glycerolphosphate was examined as follows. The reaction product of Hx -HAD hydrolase was analyzed by NMR.

The reaction product was concentrated three times by vacuum drying, appropriately diluted with D 2 O D, The 1 H-NMR spectra were acquired using a pulse sequence incorporating presaturation for water suppression. The chemical shifts of the 1 H-NMR spectra were given in ppm relative to the signals of solvents using external standards of D 2 O at 4.

The obtained NMR data was analyzed with Alice2 version 6. When measuring guanosine phosphorylase activity, cells were cultured without nucleosides to decrease background signals deriving from the added nucleosides.

Cell-free extracts were prepared as described for purifying GaR. The products R1P, R15P, RuBP, Ru1P, DHAP, and ethylene glycol were monitored by a differential refractive index detector.

Sequences of 16S ribosomal RNA genes from halophiles were obtained from the KEGG Genes database. When there were multiple 16S ribosomal RNAs in one organism, one sequence was randomly selected.

Further information on research design is available in the Nature Research Reporting Summary linked to this article. Uncropped gel images of the gel images shown in Supplementary Figs. All data for bar graphs in Fig. Artificial gene sequences encoding Ht -R15P isomerase, Hs -RbsK, Ht -RbsK, Hx -RbsK, Ht -Urdpase1, Hx -Urdpase1, Hl -Urdpase1, Hx -HAD hydrolase, Hx -FucA, and Hs -GaR are deposited in GenBank with the accession numbers LC, LC, LC, LC, LC, LC, LC, LC, LC, and LC, respectively.

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Download references. The authors are grateful to Ms. Eriko Kusaka and Mr. Haruo Fujita for NMR analysis. Step 3: Non-oxidative phase.

The ribosephosphate from step 3 is combined with another molecule of ribosephosphate to make one, carbon molecule. Excess ribosephosphate, which may not be needed for nucleotide biosynthesis, is converted into other sugars that can be used by the cell for metabolism. The carbon molecule is interconverted to create a 3-carbon molecule and a 7-carbon molecule.

The 3-carbon product can be shipped over to glycolysis if it needs. That being said, recall that we can also work our way back up to another molecule in this phase. So that 3-carbon molecule could also be shipped over from glycolysis and transformed into ribosephosphate for DNA and RNA production.

The 3-carbon molecule and the 7-carbon molecule, from the interconversion above in step 4, interconvert again to make a new 4-carbon molecule and 6-carbon molecule. The 4-carbon molecule is a precursor for amino acids, while the 6-carbon molecule can be used in glycolysis.

The same reversal of steps in option 4 can happen here as well. Overview of pentose phosphate pathway. The pentose phosphate pathway takes place in the cytosol of the cell, the same location as glycolysis. The two most important products from this process are the ribosephosphate sugar used to make DNA and RNA, and the NADPH molecules which help with building other molecules.

Non-oxidative phase:. NADPH is readily available to donate its electrons in the cell because it occurs in such high concentration. Aside from helping build molecules, what kind of benefit is this really for the cell? NADPH is able to donate its electrons to compounds that fight dangerous oxygen molecules.

Antioxidants donate electrons to neutralize dangerous oxygen radicals super reactive oxygen molecules. Once they have given away their electrons, antioxidants need to quickly reload in case there are more oxygen radicals.

NADPH is able to give antioxidants their constant flow of electrons to fight oxygen crime. Cellular respiration articles:. Glycolysis and gluconeogenesis The citric acid cycle Oxidative phosphorylation. Want to join the conversation?

Log in. Sort by: Top Voted. David Moore. Posted 8 years ago. Why does it say a 10 Carbon atom? Shouldn't it be a 10 Carbon molecule? Downvote Button navigates to signup page.

Flag Button navigates to signup page. Show preview Show formatting options Post answer. Suvradri Maitra. Posted 6 years ago. What is the significance of this pathway? Why do we have this? When does this pathway come to use? The pentose phosphate pathway is another way that the body is able to use glucose, in the form of glucose 6-phosphate.

The pathway is important because it is how our bodies create molecules for other processes such as ribose 5-phosphate being a precursor to RNA or DNA and erythrose 4-phosphate being used as an amino acid precursor. For this pathway it is important to remember that the non-oxidative stage is reversible so a molecule can enter at any point and be converted to whatever the body needs weather that be energy or materials of RNA or amino acids.

Comment Button navigates to signup page. Christos Emmanouil Loukas. Posted 4 years ago. you should update it with the enzyme names. David Grecu. Posted 7 years ago. In the "In Summary" section, under "Non-oxidative phase", it says that Ribosephosphate is also produced in the oxidative phase.

Isn't it produced only in the non-oxidative phase, whereas its precursor RibULOSEphosphate is produced in the oxidative phase? While yes, ribuloseP is the final product in oxid.

phase wrt irreversible reactions, the ribuloseP can become isomerized by an enzyme phosphopentose isomerase to become riboseP. So, really it's not a mix up. Posted 5 years ago. Is 6-phosphogluconate a carboxylic acid?

Carbon one is attached via double bond to an oxygen and a single bond to a hydroxyl molecule. Wouldn't the -oate suffix indicate an ester? Yes the form shown in the figure is actually 6-phosphogluconic acid — however at physiological pHs the acid will be deprotonated i.

Pentose phosphate pathway

The study protocol was reviewed and approved by the ethics committee of the Affiliated Cancer Hospital of Fudan University. Written informed consent forms were obtained from the patients prior to the beginning of the study.

Polymerase chain reaction PCR -amplified human TKT was cloned into pRK7-FLAG vector between Sal I and EcoR I and into pcDNA3. TKTL1 was cloned into pRK7-Flag vector between Hind III and EcoR I and into pcDNA3. CDH1 was cloned into pRK7-Flag vector between EcoR I and Hind III and into pcDNA3.

CDC20 was cloned into pRK7-Flag between Hind III and EcoR I. SKP2 and FBXW7 were cloned into pRK7-Flag by One step clone kits. The sequences of the primers used in this study are listed in Supplementary Table 1. For purifying heterodimer proteins of TKT and TKTL1, full-length TKT and TKTL1 were cloned in pET Duet3 vectors obtained from YANHUI XU Laboratory.

TKT was inserted between EcoR I and Hind III after the His-tag, while TKTL1 was cloned into the vector between Nde I and Xho I with a FLAG-tag. The antibody against for TKTL1 NBP, dilution was purchased from Novus Biologicals.

The CDC20 , dilution , SKP2 , dilution antibody was from Cell Signaling Technology. CDH1 CC43, dilution was obtained from Millipore.

The antibody against TKT sc, dilution was purchased from Santa Cruz Biotechnology. RPIA , dilution antibody was from Abcam. Anti-β-actin A, dilution , antibody was purchased from GeneScript.

Anti-Flag M, dilution , Anti-Myc M, dilution , and anti-HA M, dilution antibodies were obtained from Abmart. DAPI D was from Sigma-Aldrich.

EdU A and Azide Alexa Fluor A were purchased from Invitrogen. HEKT ATCC Number: CRL , HeLa ATCC Number: CCL-2 and MCF7 ATCC Number: HTB were purchased from Shanghai Cell Bank and tested negative for mycoplasma contamination. HeLa cells were authenticated using Short Tandem Repeat STR analysis by Shanghai Biowing Applied Biotechnology Company.

PFKFB3 knockout HeLa cell lines are kindly provided by Dr. Ye Dan, MCB laboratory, Fudan University. The guide sequence targeting the human PFKFB3 gene is 5ʹ- AGC TGA CTC GCT ACC TCA AC-3ʹ. The TKT or TKTL1-positive stable cells were lysed on ice in 0.

The precipitates were washed three times with 0. The peptides in the supernatant were collected by centrifugation and dried in a speed vacuum Eppendorf. Samples were re-dissolved in NH 4 HCO 3 buffer containing 0. Plasmid transfections were carried out by the Polyethylenimine PEI , Lipofectamine Invitrogen , or calcium phosphate methods.

In the Lipofectamine transfection method. This is important to balance the pH for transfection efficiency. The plates were swirled and placed back into the incubator. For immunoprecipitation, cells were lysed with 0. The binding complexes were washed with 0.

For immunohistochemical staining, tissue sections were deparaffinized by xylene two times and then hydrated. Sections were developed with the DAB kit, and the reaction was stopped with water.

The H-score method, which combines the immunoreactivity intensity values and the percentage of stained tumor cells, was used to quantify the positive score of each sample. Flag Beads Sigma was used for immunoprecipitation.

Ubiquitination was analyzed by immunoblotting using anti-HA and anti-Flag antibodies. CDH1 knockdown was carried out using synthetic siRNA oligonucleotides synthesized by Genepharma. A scrambled siRNA was used as a control.

For each target gene, we employed two effective target sequences to exclude off-target effects. Transfections were performed by using Lipofectamine Invitrogen.

The knockdown efficiency was verified by q-RT-PCR or western blotting. Supplementary Table 1 lists the DNA sequences for the siRNA.

To generate cells stably knocked down for TKTL1 and TKT, lentiviruses carrying pMKO empty vector, pMKO-TKTL1, or pMKO-TKT were produced in HEKT and HeLa cells, using VSVG and GAG as packaging plasmids.

Puromycin was used to select stable cells for ~5 days. The DNA sequences of the siRNAs are listed in Supplementary Table 1. For forward TKT activity measurement, we coupled measurement of enzyme activity with GAPDH enzymes. Each experiment was repeated three times.

For reverse TKT activity measurement, we measured enzyme activity through detection of either R5P for reaction 3 or E4P production for reaction 3 in reversed reactions.

Samples were added to a 1. For measuring in vivo TKT activity, cells were sonicated and centrifuged, after which the resulting supernatant was collected for analysis.

The cells were next analyzed using a fluorescence-activated cell sorter FACS. The gating strategy of flow cytometry was shown in Supplementary Fig. The wild-type TKT and TKTL1 were cloned in vector pSJ3 with 6× HIS tag at the N-terminal.

Recombinant heterodimer of TKT and TKTL1 was cloned into the pET-Duet vector. TKTL1 was fused in-frame with 6× HIS tag at the N-terminal, while TKT was inserted in the vector with a Flag tag at the C-terminal. Plasmids were transformed into E.

coli BL21 DE3 pLysS strain and protein expression was induced by addition of 0. Cells were lysed by sonication and nickel columns GE Healthcare were used to purify proteins.

Heterodimer proteins were isolated by sequential affinity purification using Nickel resin followed by FLAG beads Sigma-Aldrich.

All primers for analysis were synthesized by Generay Shanghai. Primer sequences are listed in Supplementary Table 1. The analysis was performed by using an Applied Biosystems HT Sequence Detection System, with SYBR green labeling.

The mobile phase comprised eluent A 0. The elution program was as follows, 0. The flow rate of the pump was 0.

Glyceraldehydep and dihydroxyacetone phosphate ions were monitored at precursor-product ; ribosep and xylulosep ions at ; sedulosephosphate ions at ; erythrosephosphate ions at ; fructosephosphate ions at Each measurement was obtained at least in triplicate.

The cells were washed twice with PBS after collection to completely remove labeled glucose that was not metabolized by the cells.

M2 of R5P at , IMP at , AMP at , GMP at , and PRPP at For absolute R5P concentration measurement, 0, 5, 10, 50, , , and nmol R5P from cell lysis were used to obtain an R5P standard curve.

After R5P detection via LC-MS, the absolute concentration of R5P was calculated using the R5P standard curve. Proteins were passed over the gel filtration column. The flow speed rate was 0. Fractions were collected every 0. Molecular mass was determined by Gel Filtration Calibration Kit HMW GE Healthcare.

Cells were harvested and washed with PBS twice to remove the remaining medium. DAPI was subsequently added, for nuclear staining. Results were acquired in flow cytometer or cells were observed under a fluorescence microscope.

The binding kinetics and affinity of TKT with TKTL1 protein or small molecules were analyzed by SPR Biacore T, GE Healthcare. To determine the K d of TKT and TKTL1, TKTL1 protein was diluted to a series of concentrations starting at Statistical analysis was performed using Prism 6.

Two-tailed Student's t -test was performed for the two-group analysis. Further information on research design is available in the Nature Research Reporting Summary linked to this article.

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We refrain from designating the values of these measurements as reaction rates, as the product formation rate is not necessarily constant throughout the reaction period. The Hx -RbsK recombinant protein was purified for further biochemical analyses Supplementary Fig.

Using R1P as the phosphate acceptor, the enzyme preferred ATP among NTPs Supplementary Fig. HPLC analysis of the Hx -RbsK reaction product indicated that R15P was generated from R1P Fig.

In addition, the Hx -RbsK product, R15P, was isomerized to RuBP by Ht -R15P isomerase Fig. Hx -RbsK required salt for its kinase activity, and 2. The former is designated here as Urdpase1 and the latter Urdpase2.

They can be distinguished phylogenetically Supplementary Fig. In this study, the Urdpase1 protein was investigated. Although we assumed that Urdpase1 catalyzes a nucleoside phosphorylase reaction and generates R1P, the substrate for ATP-R1PK, it was unclear which nucleosides are recognized by the enzyme.

Four Urdpase1 recombinant proteins from H. xanaduensis , H. lacusprofundi , and H. Hl -Urdpase1 and Ht -Urdpase1 were obtained as soluble proteins, while the other two formed inclusion bodies. Hl -Urdpase1 recombinant protein was purified to apparent homogeneity Supplementary Fig.

Nucleoside phosphorylase activity of purified Hl -Urdpase1 was examined toward six nucleosides, adenosine, inosine, guanosine, cytidine, uridine, and thymidine Fig. Hl -Urdpase1 exhibited highest activity toward guanosine, and could recognize adenosine and inosine to a lower extent.

Surprisingly, activity was lower at higher KCl concentrations Supplementary Fig. Kinetic analyses toward guanosine and phosphate revealed that the kinetic parameters V max and K m were 1. The identification of guanosine phosphorylase, ATP-R1PK, and R15P isomerase implied the presence of a metabolic pathway converting guanosine, phosphate, and ATP to RuBP, guanine, and ADP via R1P and R15P Fig.

Although the phosphate donor of R1P kinase is ATP, the metabolic route from guanosine to RuBP corresponds to that in the pentose bisphosphate pathway identified in Thermococcus a Nucleoside phosphorylase activity of Hl -Urdpase1 was analyzed toward six nucleosides by quantifying the released nucleobases with HPLC.

b Phosphatase activity of Hx -HAD hydrolase was investigated toward eleven sugar phosphates and pNPP by quantifying released phosphate with malachite green.

G1P glucosephosphate, G6P glucosephosphate, G16P glucose-1,6-bisphosphate, F1P fructosephosphate, F6P fructosephosphate, F16P fructose-1,6-bisphosphate, R5P ribosephosphate, Ru5P ribulosephosphate, pNPP p -nitrophenylphosphate.

c Aldolase activity of the Hx -FucA condensing DHAP and six aldehydes was examined by quantifying the residual DHAP after reactions with a coupling enzyme.

Error bars indicate standard deviations. Other than R15P isomerase, Rubisco is the sole enzyme known to utilize RuBP as a substrate. The presence of a metabolic route generating RuBP in halophilic archaea without a Rubisco suggested the presence of unidentified enzymes involved in the conversion of RuBP.

We took a comparative genomics approach, along with phylogenetic analysis, to identify enzymes that might be involved in this metabolism. We found that a gene annotated as fuculosephosphate aldolase fucA is specifically present in 17 of the 19 halophilic archaea that harbor a standalone R15P isomerase.

FucA homologs are widely distributed among the halophilic archaea and a relationship with the occurrence of R15P isomerase is not readily apparent. However, a phylogenetic analysis Supplementary Fig.

There were only two organisms H. tiamatea and Halohasta litchfieldiae that harbored a standalone R15P isomerase but not FucA.

In addition, these two genes form an operon Fig. Furthermore, in H. lacusprofundi , Halorubrum sp. PV6, and Halorubrum ezzemoulense , the four genes encoding ATP-R1PK, R15P isomerase, HAD hydrolase, and FucA form an operon.

The two genes fucA and hadhlase display a stark co-occurrence with the three genes encoding guanosine phosphorylase, ATP-R1PK, and R15P isomerase, strongly raising the possibilities that these five proteins are metabolically related.

We examined the possibility that the proteins annotated as HAD hydrolase and FucA are involved in RuBP metabolism. The HAD superfamily hydrolase from Arabidopsis thaliana is known to catalyze the phosphatase reaction of ribose 5-phosphate R5P to ribose Supplementary Fig.

Based on the structural similarity between R5P and RuBP Supplementary Fig. The gene encoding HAD hydrolase from H. coli and the recombinant protein Hx -HAD hydrolase was purified to apparent homogeneity Supplementary Fig.

Phosphatase activity was measured toward 12 sugar phosphates. We observed notable phosphate production only when RuBP was used as a substrate Fig. The optimal KCl concentration was higher than 2. Kinetic analysis toward RuBP Supplementary Fig.

We designate the enzyme as RuBP phosphatase. Dephosphorylation of RuBP can result in the generation of three reaction products, ribulosephosphate Ru1P , ribulosephosphate Ru5P , and ribulose.

We thus carried out an 1 H-NMR analysis of the reaction product Supplementary Fig. Although chemical shifts derived from RuBP were also detected, the chemical shifts specific to a cyclized Ru1P shown in a previous report 35 were confirmed in the reaction product Supplementary Fig.

This suggested that RuBP phosphatase catalyzes the hydrolysis of the phosphate group at the C5-position of RuBP, generating Ru1P Fig. Classical FucA is an enzyme catalyzing the aldolase reaction of fuculosephosphate Fu1P. Fu1P is cleaved into dihydroxyacetone phosphate DHAP and l -lactaldehyde Supplementary Fig.

We noted that the chemical structures of Ru1P and Fu1P are similar and the difference lies only in the C6 methyl group Supplementary Fig. coli , which displays xanaduensis Hx -FucA , could recognize DHAP and glycolaldehyde Recombinant Hx -FucA was thus prepared using E.

coli and purified to apparent homogeneity Supplementary Fig. As Ru1P is not commercially available, we examined whether Hx -FucA can catalyze the aldolase reaction condensing DHAP and aldehydes by quantifying the residual DHAP.

The aldehydes tested were acetaldehyde, propionaldehyde, isobutylaldehyde, dl -glyceraldehyde, dl -lactaldehyde, and glycolaldehyde Supplementary Fig. The protein displayed aldolase activity toward DHAP and all tested aldehydes Fig. The result implied that although the substrate specificity of the enzyme was broad, it could catalyze the aldolase reaction cleaving Ru1P to DHAP and glycolaldehyde.

With HPLC, we further confirmed that the reaction product of the Hx -FucA reaction with DHAP and glycolaldehyde displayed an elution time identical to that of the RuBP phosphatase reaction product Supplementary Fig.

An increase in activity was observed with the addition of ZnCl 2 to the reaction mixture, while a decrease was observed with addition of EDTA, suggesting that the enzyme was dependent on zinc cations Supplementary Fig.

In addition, we examined whether Hx -RuBP phosphatase and the Hx -FucA protein together could generate DHAP from RuBP. Only when both proteins were present in the reaction mixture, DHAP was produced from RuBP Supplementary Fig. Furthermore, kinetic analyses of the enzyme revealed that V max and K m toward glycolaldehyde were 2.

Based on the results obtained here, we propose that the standalone R15P isomerase is a component of a previously unidentified nucleoside metabolic pathway converting the ribose moiety of guanosine to glycolaldehyde and DHAP Fig.

As the pathway does not involve Rubisco, we here designate this pathway the non-carboxylating pentose bisphosphate pathway. Although DHAP can be metabolized by central sugar metabolism, the fate of glycolaldehyde was still unclear. However, we could not identify a candidate enzyme based on genome information that would metabolize glycolaldehyde.

Upon measuring enzyme activity in the H. salinarum cell-free extract that could potentially convert glycolaldehyde, we were able to detect reducing activity on glycolaldehyde using NADH as the electron donor. From the cell-free extract of H. salinarum cultured with nucleosides, we purified the protein displaying glycolaldehyde reductase activity to apparent homogeneity Supplementary Fig.

Kinetic analysis toward glycolaldehyde Supplementary Fig. Taking into account the original annotation, we further examined whether the protein catalyzes the sn -glycerolphosphate dehydrogenase reaction or not Supplementary Fig.

The purified enzyme did not display notable levels of activity for the oxidation of glycerolphosphate nor the reduction of DHAP. coli , and partially purified Supplementary Fig.

As expected, the recombinant protein Hs -GaR displayed high levels of glycolaldehyde reductase activity As sn -glycerolphosphate dehydrogenase is presumed to contribute to the biosynthesis of archaeal membrane lipid precursors by reducing DHAP to glycerolphosphate, the enzyme would seem to be essential in all archaea.

However, we found another gene annotated as sn -glycerolphosphate dehydrogenase distributed in all halophilic archaea including H. salinarum is encoded on a plasmid. Although half possess a complete set of genes forming the non-carboxylating pentose bisphosphate pathway, the other half does not.

The biochemical analyses described above involves enzymes from different species of halophilic archaea. In order to confirm that the proposed pathway is present in a single species, we examined the enzyme activities in the cell-free extract of H. When each substrate was added to the reaction mixture including cell-free extracts, the products of guanosine phosphorylase R1P , ATP-dependent R1P kinase R15P , RuBP phosphatase Ru1P , and glycolaldehyde reductase ethylene glycol reactions were detected Supplementary Fig.

On the other hand, when R15P was added to the reaction mixture, we could not detect RuBP, the product of R15P isomerase. However, instead of RuBP, we could observe the generation of Ru1P Supplementary Fig.

This result implied that the RuBP generated from R15P by R15P isomerase was subsequently converted to Ru1P by RuBP phosphatase. To confirm Ru1P aldolase activity, as Ru1P is not commercially available, a coupling reaction catalyzed by RuBP phosphatase and Ru1P aldolase was examined.

As a result, DHAP, which is presumed to be produced by Ru1P aldolase from Ru1P, was clearly detected when RuBP and ZnCl 2 were added Supplementary Fig.

The predicted activities of the enzymes encoded by the six genes were detectable in H. salinarum , suggesting the presence of the non-carboxylating pentose bisphosphate pathway in this organism. Based on the results of this study, we propose a previously unrecognized nucleoside degradation pathway, the non-carboxylating pentose bisphosphate pathway, in halophilic archaea Fig.

Although the metabolism from guanosine to RuBP via R15P is similar to that in the pentose bisphosphate pathway in Thermococcales, the downstream route is unique. We can assume that the physiological role of the pathway is to convert the ribose moiety of nucleoside s to DHAP and ethylene glycol.

DHAP can be utilized in various metabolisms, including oxidation to pyruvate via glyceraldehyde 3-phosphate, gluconeogenesis, and conversion to glycerol for utilization in membrane lipid biosynthesis and osmolyte production. On the other hand, the metabolic fate of ethylene glycol is still unclear and further examination will be necessary to understand if and how the cells utilize the two carbons deriving from pentoses.

Our results and the distribution of gene homologs suggest the presence of multiple variations of nucleoside degradation pathways in halophilic archaea, all involving the pentose bisphosphates R15P and RuBP.

One common feature in the nucleoside degradation pathways found in halophilic archaea is that the ADP-R1PK found in Thermococcales is replaced by the ATP-R1PK identified in this study Figs. As shown in Table 1 , among the 63 species of halophilic archaea whose genome sequences have been determined, 44 species harbor R15P isomerase homologs on their genomes, suggesting the presence of metabolism involving the pentose bisphosphates R15P and RuBP in these organisms.

Among these, 25 species seem to utilize Rubisco for the metabolism of RuBP. The non-carboxylating pentose bisphosphate pathway identified in this study, utilizing RuBP phosphatase and Ru1P aldolase, is found in 17 halophile species and is also widely distributed.

As for NMP phosphorylase, homologs are only found in species with a Rubisco. Among the 25 species with Rubisco, 14 harbor an NMP phosphorylase homolog, while 11 do not. It thus seems that there are three major variations of the pentose bisphosphate pathway in halophiles that account for 42 of the 63 species shown in Table 1 ; i one with Rubisco and NMP phosphorylase, as seen in members of Thermococcales Fig.

The distribution of these variations among the halophilic archaea is not linked to the phylogenetic relationships of their source organisms Fig.

This raises the possibility that there may be even more variations of nucleoside degradation pathways in halophilic archaea that involve pentose bisphosphates. The results obtained in this study and the distribution of relevant genes imply the occurrence of three types of nucleoside degradation pathways in halophilic archaea.

The three metabolic routes are with NMP phosphorylase and Rubisco a , with Rubisco but without NMP phosphorylase b , without Rubisco or NMP phosphorylase but with RuBP phosphatase and Ru1P aldolase c.

The phylogenetic tree was constructed using 16S rRNA gene sequences. A sequence from Thermoplasma volcanium tvo was used as an outgroup. The colors of the circles correspond to those of the nucleoside metabolic pathways in Fig. Organisms shown in red and pink codes indicate the halophilic archaea shown in Fig.

The organisms in red codes show those whose proteins were actually examined in this study. Among them, Halorhabdus utahensis and Halorhabdus tiamatea harbor homologs of the classical NOPP pathway found in bacteria and eukaryotes, and the NOPP pathway may be responsible for nucleoside degradation in these species.

However, there are still 19 halophilic archaea whose genome sequences do not provide any clues as to how they carry out nucleoside degradation. The homolog tends to occur in these halophiles, including 9 species in Haloarcula , Haloplanus , and Halohasta , and might provide clues to identify additional pathways involved in nucleoside or pentose metabolism.

Analysis of recombinant Hl -Urdpase1 indicated that the enzyme preferred guanosine as the nucleoside substrate for the phosphorylase reaction.

In addition to the guanosine phosphorylase, almost all halophilic archaea harbor a second uridine phosphorylase homolog, Urdpase2. Although these second homologs do not form an operon with genes of the pentose bisphosphate pathway, there is a high possibility that the gene products also generate R1P via nucleoside phosphorolysis.

Phylogenetic analysis clearly revealed that Urdpase1 and Urdpase2 are distinct Supplementary Fig. Identifying the nucleoside substrate of Urdpase2 will add to our understanding of nucleoside degradation in halophilic archaea. It is intriguing that only one of the two nucleoside phosphorylase genes, the guanosine phosphorylase gene, forms an operon with the genes of the non-carboxylating pentose bisphosphate pathway.

This may be related to the high GC contents in genomes of halophilic archaea such as H. salinarum Two R1P kinases have been identified in previous studies; an ADP-dependent R1P kinase from the hyperthermophilic archaeon T.

When comparing the properties of the three enzymes, the R1P kinase from T. kodakarensis is ADP-dependent, whereas the enzymes from P.

calidifontis and H. salinarum are ATP-dependent. On the other hand, the enzymes from T. kodakarensis and H. xanaduensis display strict substrate specificity toward R1P, while the R1P kinase from P. calidifontis can recognize cytidine and uridine in addition to R1P Neither R15P isomerase nor NMP phosphorylase gene homologs are present in P.

calidifontis , resembling the case of the ten halophilic archaea with only nucleoside phosphorylase and ATP-R1PK homologs described above. On the other hand, the presence of R1P kinase in E. coli has been suggested A protein encoded by phnN was identified that displays kinase activity towards R15P, leading to the production of PRPP.

A protein responsible for the proposed second reaction, although unidentified, would be an R1P kinase. coli possesses several enzymes displaying similarity with the archaeal proteins that display R1P kinase activity.

As these include proteins whose functions have not been determined, one of them may also be R1P kinase. R1P kinase and its product, R15P, may be distributed through Archaea and Bacteria more widely than previously expected.

Unless mentioned otherwise, chemical reagents were purchased from Nacalai Tesque Kyoto, Japan , FUJIFILM Wako Pure Chemicals Osaka, Japan , or Merck Darmstadt, Germany. Strains and plasmids used in this study are listed in Supplementary Table 2.

When necessary, nucleosides were added into the medium. Escherichia coli DH5α strain used for plasmid construction and E. Plasmids to express genes encoding Hs -R15P isomerase and Hs -Urdpase1 were constructed as follows. Coding regions of Hs -R15P isomerase and Hs -Urdpase1 genes were amplified by PCR using genomic DNA from H.

Sequences of these primers are listed in Supplementary Table 3. The resulting expression plasmids are designated pET-Hs-R15P isomerase and pET-Hs-Urdpase1 Supplementary Table 2.

Expression plasmids for genes encoding Ht -R15P isomerase, Hs -RbsK, Ht -RbsK, Hx -RbsK, Ht -Urdpase1, Hx -Urdpase1, Hl -Urdpase1, Hx -HAD hydrolase, Hx -FucA, and Hs -GaR Supplementary Table 2 were prepared as follows.

Genes were designed and synthesized Integrated DNA Technologies, Coralville, IA to decrease their GC contents, and optimize their codons to enhance gene expression in E.

For all 10 resulting expression plasmids Supplementary Table 2 with the two plasmids described above, we carried out DNA sequencing analysis and confirmed the absence of unintended mutation. The designed sequences of each gene are shown in Supplementary Fig.

The constructed expression plasmids were introduced into E. coli Rosetta DE3 for genes encoding R15P isomerase, RbsK, HAD hydrolase, and FucA or E. coli BLCodonPlus DE3 -RIL for genes encoding Urdpase1 and GaR. Proteins were eluted with a linear gradient of NaCl 0 to 1. The same buffer was used as mobile phase.

Hl -Urdpase1 and Hx -HAD hydrolase recombinant proteins were purified with Bio-Scale CHT and Superdex After centrifugation, the supernatants were applied to Bio-Scale CHT equilibrated with the same buffers used for cell suspension.

The concentrations of the purified enzymes were determined with the Protein Assay System Bio-Rad , using bovine serum albumin BSA Thermo Fisher Scientific, Waltham, MA as a standard. Protein homogeneity was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis SDS-PAGE.

After cell density OD reached 0. Fractions displaying glycolaldehyde reductase activity were collected. The same buffer was used as a mobile phase. Ribose 1-phosphate can interact spontaneously with ATP resulting in a release of hydrogen ion, ADP and a ribose 1,5-biphosphate.

Ribose 1,5-biphosphate is then phosphorylated through a ribose 1,5-bisphosphokinase resulting in the release of ADP and phosphoribosyl pyrophosphate. Phosphoribosyl pyrophosphate will then participate in the purine nucleotides de novo biosynthesis pathway.

Alternatively pentose phosphate and D-ribose 5-phosphate's interaction can be phosphorylated through an ATP driven ribose-phosphate diphosphokinase resulting in a release of a hydrogen ion, an AMP and a phosphoribosyl pyrophosphate which will again participate in the purine nucleotides de novo biosynthesis pathway.

PRPP Biosynthesis References Hove-Jensen B, Nygaard P: Phosphoribosylpyrophosphate synthetase of Escherichia coli, Identification of a mutant enzyme. Eur J Biochem. Hove-Jensen B, Harlow KW, King CJ, Switzer RL: Phosphoribosylpyrophosphate synthetase of Escherichia coli.

Properties of the purified enzyme and primary structure of the prs gene. J Biol Chem. Hove-Jensen B, Rosenkrantz TJ, Haldimann A, Wanner BL: Escherichia coli phnN, encoding ribose 1,5-bisphosphokinase activity phosphoribosyl diphosphate forming : dual role in phosphonate degradation and NAD biosynthesis pathways.

J Bacteriol.

How does it happen? Biosyntheis CAS PubMed Boisynthesis Scholar Watson, G. That being Sports nutrition tips and tricks, recall that we Ribose biosynthesis pathway also work pathwwy way back up to another molecule in this Ribose biosynthesis pathway. With HPLC, we further confirmed that biosyynthesis reaction product of the Hx -FucA reaction with DHAP and glycolaldehyde displayed an elution time identical to that of the RuBP phosphatase reaction product Supplementary Fig. TKTL1 knockdown not only lowered the levels of R5P-containing molecules, but also abolished the effects of CDH1 overexpression in HEKT cells Fig. In late G 1 and S phases, transketolase-like 1 TKTL1 is overexpressed and forms stable TKTL1-transketolase heterodimers that accumulate ribosephosphate. Download references. Overview of pentose phosphate pathway.

Ribose biosynthesis pathway -

Show In Black and White Clear. Adenosine diphosphate Adenosine monophosphate Adenosine triphosphate D-Ribose 5-phosphate Hydrogen Ion Magnesium Phosphate Phosphoribosyl pyrophosphate Ribose 1,5-bisphosphate Ribose 1-phosphate.

ATP-binding protein phnN Phosphopentomutase Ribose-phosphate pyrophosphokinase. Enter relative concentration values without units. For the best results, view the pathway in Black and White. SVG Image Simple SVG Image Large Font SVG Image Simple Large Font SVG Image Greyscale SVG Image BioPAX SBGN SBML PWML PNG Image All files.

Turn On Complex Membranes Show Simplified Version Show Large Font Version Show Simplified Large Font Version Set Background White. SVG Image. Simple SVG Image. Large Font SVG Image. Simple Large Font SVG Image. The increase in glucose metabolism is accompanied not only by an increase of lactate production but also by an increase in pentose production for RNA and DNA synthesis Eigenbrodt et al.

However, the importance of the pentose phosphate pathway in tumours has been overlooked. These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves. This is a preview of subscription content, log in via an institution.

Unable to display preview. Download preview PDF. Basu, T. Article PubMed CAS Google Scholar. Ben-Yoseph, O. Boros, L. et al. PubMed CAS Google Scholar. Cascante, M.

Champe, P. Oxidative reactions and nonoxidative reactions, pp. Google Scholar. Cornish-Bowden, A. Rehm, H. Chapter Google Scholar.

Eigenbrodt, E. Beitner, R. Fell, D. Georgiannos, S. Cancer 68, — Groen, A. l01— in Control of Metabolic Processes ed. Heinrich, R. Henquin, N. Hiatt, H. The pathway of nucleic ribose synthesis in human carcinoma cell in tissue culture J.

Horecker, B. Kacser, H. Given the structural and functional importance of DNA and RNA for all living things, there are many layers of quality control to help avoid and correct mistakes when DNA and RNA are initially made. While the products of glycolysis are sent through the rest of cellular respiration to produce energy see video about glycolysis here , there is also an alternative branch off glycolysis to produce the sugars that make up DNA and RNA.

This pathway, called the Pentose Phosphate Pathway, is special because no energy in the form of ATP, or adenosine triphosphate, is produced or used up in this pathway. Similarly to some of the processes in cellular respiration, the molecules that go through the pentose phosphate pathway are mostly made of carbon.

The easiest way to understand this pathway is to follow the carbon. The breakdown of the simple sugar, glucose, in glycolysis provides the first 6-carbon molecule required for the pentose phosphate pathway.

During the first step of glycolysis, glucose is transformed by the addition of a phosphate group, generating glucosephosphate, another 6-carbon molecule. The pentose phosphate pathway can use any available molecules of glucosephosphate, whether they are produced by glycolysis or other methods.

Cellular respiration overview. Now, we are ready to enter the first of two phases of the pentose phosphate pathway: 1 The oxidative phase and 2 The non-oxidative phase.

Oxidation is the breakdown of a molecule as it loses at least one of its electrons. This phase is made up of 2 irreversible steps:. Step Glucosephosphate is oxidized to form lactone. Following the oxidation of glucosephosphate, another reaction, catalyzed by a different enzyme, uses water to form 6-phosphogluconate, the linear product.

NADPH is similar in structure and function as the high energy electron shuttle, NADH, mentioned in the cellular respiration articles.

NADPH has an added phosphate group and is used in the cell to donate its electrons, just like NADH. NADPH is often used in reactions that build molecules and occurs in a high concentration in the cell, so that it is readily available for these types of reactions.

Step 1: Oxidative phase. This new 5-carbon molecule is called ribulosephosphate. Step 2: Oxidative phase. The non-oxidative phase is really handy because these reactions are reversible.

This allows different molecules to enter the pentose phosphate pathway in different areas of the non-oxidative phase and be transformed up until the first molecule of the non-oxidative phase ribulosephosphate.

Ribulosephosphate is the precursor to the sugar that makes up DNA and RNA, and is also a product of the oxidative stage. Ribulose phosphate can be converted into two different 5-carbon molecules. One is the sugar used to make up DNA and RNA called, ribose phosphate and this is the molecule we will focus on.

Step 3: Non-oxidative phase. The ribosephosphate from step 3 is combined with another molecule of ribosephosphate to make one, carbon molecule. Excess ribosephosphate, which may not be needed for nucleotide biosynthesis, is converted into other sugars that can be used by the cell for metabolism.

The carbon molecule is interconverted to create a 3-carbon molecule and a 7-carbon molecule. The 3-carbon product can be shipped over to glycolysis if it needs. That being said, recall that we can also work our way back up to another molecule in this phase.

So that 3-carbon molecule could also be shipped over from glycolysis and transformed into ribosephosphate for DNA and RNA production. The 3-carbon molecule and the 7-carbon molecule, from the interconversion above in step 4, interconvert again to make a new 4-carbon molecule and 6-carbon molecule.

The 4-carbon molecule is a precursor for amino acids, while the 6-carbon molecule can be used in glycolysis. The same reversal of steps in option 4 can happen here as well. Overview of pentose phosphate pathway.

PATHWAY: ptahway Help Entry rn Pathway Name Pentose Ribose biosynthesis pathway pathway. The pentose phosphate pathway is a process of Rbose turnover pathawy produces NADPH as reducing equivalents and pentoses as essential parts of Ribose biosynthesis pathway. There Energy-packed meal ideas two different phases in the pathway. One is irreversible oxidative phase in which glucose-6P is converted to ribulose-5P by oxidative decarboxylation, and NADPH is generated [MD: M ]. The other is reversible non-oxidative phase in which phosphorylated sugars are interconverted to generate xylulose-5P, ribulose-5P, and ribose-5P [MD: M ]. This pathway map also shows the Entner-Doudoroff pathway where 6-P-gluconate is dehydrated and then cleaved into pyruvate and glyceraldehyde-3P [MD: M ]. Immune-boosting self-care practices you're seeing this message, it means we're Biosynthess trouble loading external resources biiosynthesis our website. org are unblocked. To log in and use all the features of Khan Academy, please enable JavaScript in your browser. Get AI Tutoring NEW. Search for courses, skills, and videos. Carbohydrate Metabolism.

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