Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.2.1.13 (glyceraldehyde-3-phosphate dehydrogenase)
6,511 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Thiol-disulfide bond balance is generally maintained in bacteria by thioredoxin reductase-thioredoxin and/or glutathione-glutaredoxin systems. Some gram-positive bacteria, including Lactococcus lactis, do not produce glutathione, and the thioredoxin system is presumed to be essential. We constructed an L. lactis trxB1 mutant. The mutant was obtained under anaerobic conditions in the presence of dithiothreitol (DTT). Unexpectedly, the trxB1 mutant was viable without DTT and under aerated static conditions, thus disproving the essentiality of this system. Aerobic growth of the trxB1 mutant did not require glutathione, also ruling out the need for this redox maintenance system. Proteomic analyses showed that known oxidative stress defense proteins are induced in the trxB1 mutant. Two additional effects of trxB1 were not previously reported in other bacteria: (i) induction of proteins involved in fatty acid or menaquinone biosynthesis, indicating that membrane synthesis is part of the cellular response to a redox imbalance, and (ii) alteration of the isoforms of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GapB). We determined that the two GapB isoforms in L. lactis differed by the oxidation state of catalytic-site cysteine C152. Unexpectedly, a decrease specific to the oxidized, inactive form was observed in the trxB1 mutant, possibly because of proteolysis of oxidized GapB. This study showed that thioredoxin reductase is not essential in L. lactis and that its inactivation triggers induction of several mechanisms acting at the membrane and metabolic levels. The existence of a novel redox function that compensates for trxB1 deficiency is suggested.
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PMID:Roles of thioredoxin reductase during the aerobic life of Lactococcus lactis. 1562 31

In Bacillus subtilis, the NADPH-dependent glyceraldehyde-3-phosphate dehydrogenase (GapB) and the phosphoenolpyruvate carboxykinase (PckA) enzymes are necessary for efficient gluconeogenesis from Krebs cycle intermediates. gapB and pckA transcription is repressed in the presence of glucose but not via CcpA, the major transcriptional regulator for catabolite repression in B. subtilis. A B. subtilis mini-Tn10 transposant library was screened for clones affected in catabolite repression of gapB. Inactivation of a previously unknown gene, yqzB (renamed ccpN for control catabolite protein of gluconeogenic genes), was found to relieve not only gapB but also pckA transcription from catabolite repression. Purified CcpN specifically bound to the gapB and pckA promoters. ccpN is co-transcribed constitutively with another unknown gene, yqfL. A yqfL deletion lowers the level of gapB and pckA transcription threefold under both glycolytic and gluconeogenic conditions and a ccpN deletion is epistatic over a yqfL deletion. YqfL is thus a positive regulator of the expression of gapB and pckA, the effect of which is not influenced by the metabolic regime of the cell but appears to be mediated by CcpN. ccpN has homologues in many Firmicutes, but not all, while yqfL homologues are widely distributed in Eubacteria and also present in some plants. In all analysed bacterial genomes, ccpN and yqfL are physically linked together or to putative gluconeogenic genes. CcpN thus orchestrates a novel CcpA-independent mechanism for catabolite repression of gluconeogenic genes highly conserved in Firmicutes and appears as a functional analogue of FruR in Enterobacteria. The physiological significance of the regulation mediated via the three B. subtilis global transcription regulators, CcpA, CggR and CcpN, is discussed.
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PMID:CcpN (YqzB), a novel regulator for CcpA-independent catabolite repression of Bacillus subtilis gluconeogenic genes. 1572 May 52

Corynebacterium glutamicum gapA and gapB encode glyceraldehyde-3-phosphate dehydrogenases (GAPDHs) that differ in molecular weight and activity in the presence of ATP. Comparative genome analysis revealed that GapA, the product of gapA, represented the canonical GAPDH that is highly conserved across the three major life forms. GapB, with an additional 110-residue-long sequence upstream of its GAPDH-specific domain, was homologous only to select microbial putative GAPDHs. Upon gene disruption, the initial growth rates of the wild-type, DeltagapA and DeltagapB strains on glucose (0.77, 0.00 and 0.76 h(-1), respectively), lactate (0.20, 0.18 and 0.15 h(-1), respectively), pyruvate (0.39, 0.29 and 0.20 h(-1), respectively), and acetate (0.06, 0.06 and 0.04 h(-1), respectively), implied that GapA was indispensable for growth on glucose, that GapB, but not GapA, affected early growth on acetate, and that GapB had a greater influence on growth under gluconeogenic conditions than GapA. The disruption of either gapA or gapB showed no significant effect on the transcription of any of the other gap cluster genes although it led to reduced triosephosphate isomerase (TPI) activities. Glycolytic GAPDH activity at low in vitro ATP concentrations was solely attributed to the 35.9-kDa GapA. At higher ATP concentrations, the same activity was attributed to the 51.2-kDa GapB. Both enzymes, however, exhibited similar NADP-dependent GAPDH activities at the higher ATP concentrations. In effect therefore, the GAPDH-catalyzed reaction at low ATP concentrations was irreversible, with all the glycolytic activity strictly NAD-dependent and attributed to GapA. At higher ATP concentrations, the reaction was reversible, with glycolytic activity NAD- or NADP-dependent and attributed to GapB, while gluconeogenic activity was attributable to both GapA and GapB.
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PMID:Corynebacterium glutamicum glyceraldehyde-3-phosphate dehydrogenase isoforms with opposite, ATP-dependent regulation. 1592

When tobacco (Nicotiana tabacum) plants were transferred from the dark to continuous white light, the steady-state mRNA levels transcribed from the nuclear genes encoding chloroplast (GapA and GapB) glyceraldehyde-3-phosphate dehydrogenase increased at least 30- to 50-fold, while the mRNA level for the cytosolic enzyme (GapC) increased only 10-fold. Kinetic analyses show that the rates of mRNA accumulation for GapA and GapB are identical reaching steady-state levels after 24-48 h in light. In contrast, mRNA accumulation for the GapC gene shows a completely different kinetic pattern, accumulating much faster than that of GapA and GapB. These results suggest that expression of GapC and GapA/B genes are controlled by different light regulated mechanisms and nuclear run-on analyses suggest that these effects are primarily due to increased transcription.
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PMID:Differential light regulated expression of nuclear genes encoding chloroplast and cytosolic glyceraldehyde-3-phosphate dehydrogenase in Nicotiana tabacum. 1645 34

To identify novel targets for metabolic engineering of riboflavin production, we generated about 10,000 random, transposon-tagged mutants of an industrial, riboflavin-producing strain of Bacillus subtilis. Process-relevant screening conditions were established by developing a 96-deep-well plate method with raffinose as the carbon source, which mimics, to some extent, carbon limitation in fed batch cultures. Screening in raffinose and complex LB medium identified more efficiently riboflavin overproducing and underproducing mutants, respectively. As expected for a "loss of function" analysis, most identified mutants were underproducers. Insertion mutants in two genes with yet unknown function, however, were found to attain significantly improved riboflavin titers and yields. These genes and possibly further ones that are related to them are promising candidates for metabolic engineering. While causal links to riboflavin production were not obvious for most underproducers, we demonstrated for the gluconeogenic glyceraldehyde-3-phosphate dehydrogenase GapB how a novel, non-obvious metabolic engineering strategy can be derived from such underproduction mutations. Specifically, we improved riboflavin production on various substrates significantly by deregulating expression of the gluconeogenic genes gapB and pckA through knockout of their genetic repressor CcpN. This improvement was also verified under the more process-relevant conditions of a glucose-limited fed-batch culture.
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PMID:Screening of Bacillus subtilis transposon mutants with altered riboflavin production. 1858 93

A new role is reported for CP12, a highly unfolded and flexible protein, mainly known for its redox function with A(4) glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Both reduced and oxidized CP12 can prevent the in vitro thermal inactivation and aggregation of GAPDH from Chlamydomonas reinhardtii. This mechanism is thus not redox-dependent. The protection is specific to CP12, because other proteins, such as bovine serum albumin, thioredoxin, and a general chaperone, Hsp33, do not fully prevent denaturation of GAPDH. Furthermore, CP12 acts as a specific chaperone, since it does not protect other proteins, such as catalase, alcohol dehydrogenase, or lysozyme. The interaction between CP12 and GAPDH is necessary to prevent the aggregation and inactivation, since the mutant C66S that does not form any complex with GAPDH cannot accomplish this protection. Unlike the C66S mutant, the C23S mutant that lacks the N-terminal bridge is partially able to protect and to slow down the inactivation and aggregation. Tryptic digestion coupled to mass spectrometry confirmed that the S-loop of GAPDH is the interaction site with CP12. Thus, CP12 not only has a redox function but also behaves as a specific "chaperone-like protein" for GAPDH, although a stable and not transitory interaction is observed. This new function of CP12 may explain why it is also present in complexes involving A(2)B(2) GAPDHs that possess a regulatory C-terminal extension (GapB subunit) and therefore do not require CP12 to be redox-regulated.
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PMID:CP12 from Chlamydomonas reinhardtii, a permanent specific "chaperone-like" protein of glyceraldehyde-3-phosphate dehydrogenase. 1928 2

The Gram-positive bacterium Staphylococcus aureus contains two glyceraldehyde-3-phosphate dehydrogenase (GAPDH) homologues known as GapA and GapB. GapA has been characterized as a functional GAPDH protein, but currently there is no biological evidence for the role of GapB in metabolism in S. aureus. In this study we show through a number of complementary methods that S. aureus GapA is essential for glycolysis while GapB is essential in gluconeogenesis. These proteins are reciprocally regulated in response to glucose concentrations, and both are influenced by the glycolysis regulator protein GapR, which is the first demonstration of the role of this regulator in S. aureus and the first indication that GapR homologues control genes other than those within the glycolytic operon. Furthermore, we show that both GapA and GapB are important in the pathogenesis of S. aureus in a Galleria mellonella model of infection, showing for the first time in any bacteria that both glycolysis and gluconeogenesis have important roles in virulence.
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PMID:Comparison of the regulation, metabolic functions, and roles in virulence of the glyceraldehyde-3-phosphate dehydrogenase homologues gapA and gapB in Staphylococcus aureus. 2087 89

In Chlamydomonas reinhardtii, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) consists of four GapA subunits. This A4 GAPDH is not autonomously regulated, as the regulatory cysteine residues present on GapB subunits are missing in GapA subunits. The regulation of A4 GAPDH is provided by another protein, CP12. To determine the molecular mechanisms of regulation of A4 GAPDH, we mutated three residues (R82, R190, and S195) of GAPDH of C. reinhardtii. Kinetic studies of GAPDH mutants showed the importance of residue R82 in the specificity of GAPDH for NADPH, as previously shown for the spinach enzyme. The cofactor NADPH was not stabilized through the 2'-phosphate by the serine 195 residue of the algal GAPDH, unlike the case in spinach. The mutation of R190 also led to a structural change that was not observed in the spinach enzyme. This mutation led to a loss of activity for NADPH and NADH, indicating the crucial role of this residue in maintaining the algal GAPDH structure. Finally, the interaction between GAPDH mutants and wild-type and mutated CP12 was analyzed by immunoblotting experiments, surface plasmon resonance, and kinetic studies. The results obtained with these approaches highlight the involvement of the last residue of CP12, Asp80, in modulating the activity of GAPDH by preventing access of the cofactor NADPH to the active site. These results help us to bridge the gap between our knowledge of structure and our understanding of functional biology in GAPDH regulation.
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PMID:Molecular mechanism of NADPH-glyceraldehyde-3-phosphate dehydrogenase regulation through the C-terminus of CP12 in Chlamydomonas reinhardtii. 2136 64

Enzymatic synthesis of some industrially important compounds depends heavily on cofactor NADPH as the reducing agent. This is especially true in the synthesis of chiral compounds that are often used as pharmaceutical intermediates to generate the correct stereochemistry in bioactive products. The high cost and technical difficulty of cofactor regeneration often pose a challenge for such biocatalytic reactions. In this study, to increase NADPH bioavailability, the native NAD(+)-dependent glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gapA gene in Escherichia coli was replaced with a NADP(+)-dependent gapB from Bacillus subtilis. To overcome the limitation of NADP(+) availability, E. coli NAD kinase, nadK was also coexpressed with gapB. The recombinant strains were then tested in three reporting systems: biosynthesis of lycopene, oxidation of cyclohexanone with cyclohexanone monooxygenase (CHMO), and an anaerobic system utilizing 2-haloacrylate reductase (CAA43). In all the reporting systems, replacing NAD(+)-dependent GapA activity with NADP(+)-dependent GapB activity increased the synthesis of NADPH-dependent compounds. The increase was more pronounced when NAD kinase was also overexpressed in the case of the one-step reaction catalyzed by CAA43 which approximately doubled the product yield. These results validate this novel approach to improve NADPH bioavailability in E. coli and suggest that the strategy can be applied in E. coli or other bacterium-based production of NADPH-dependent compounds.
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PMID:Improvement of NADPH bioavailability in Escherichia coli by replacing NAD(+)-dependent glyceraldehyde-3-phosphate dehydrogenase GapA with NADP (+)-dependent GapB from Bacillus subtilis and addition of NAD kinase. 2404 43


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