Molecular characterization of the Escherichia coli K-12 zwf gene encoding glucose 6-phosphate dehydrogenase

Rowley, Daniel L.; Wolf, Richard M.

doi:10.1128/jb.173.3.968-977.1991

Cited by 90 publications

(85 citation statements)

References 57 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Amino acid sequence around Lys-21 in Leuconostoc mesenteroides G6PD and homologous sequences in other G6PDs. The sequences are from human (H) (Persico et al, 1986), rat (R) ( Ho et al, 1988), Drosophila melanogaster (D) (Fouts et al, 1988), Saccharomyces cerevisiae (S) (Nogae & Johnston, 1990;Thomas et al, 1991), Escherichia coli (E) (Rowley & Wolf, 1991), Zymomonas mobilis (Z) (Barne11 et al, 1990), and L. mesenteroides (L) . The numbers in parentheses indicate the number of the first amino acid in each sequence.…”

Section: Methodsmentioning

confidence: 99%

Lysine‐21 of Leuconostoc mesenteroides glucose 6‐phosphate dehydrogenase participates in substrate binding through charge–charge interaction

Lee

Levy

1992

Protein Science

View full text Add to dashboard Cite

Leuconostoc mesenteroides glucose 6-phosphate dehydrogenase (G6PD) was isolated in high yield and purified to homogeneity from a newly constructed strain of Escherichia coli which lacks its own glucose 6-phosphate dehydrogenase gene. Lys-21 is one of two lysyl residues in the enzyme previously modified by the affinity labels pyridoxal 5"phosphate and pyridoxal 5'-diphosphate-5'-adenosine, which are competitive inhibitors of the enzyme with respect to glucose 6-phosphate (LaDine, J.R., Carlow, D., Lee, W.T., Cross, R.L., Flynn, T.G., & Levy, H.R., 1991, J. Biol. Chem. 266, 5558-5562). K21R and K2lQ mutants of the enzyme were purified to homogeneity and characterized kinetically to determine the function of Lys-21. Both mutant enzymes showed increased K,-values for glucose 6-phosphate compared to wild-type enzyme: 1.4-fold (NAD-linked reaction) and 2.1-fold (NADP-linked reaction) for the K21R enzyme, and 36-fold (NAD-linked reaction) and 53-fold (NADP-linked reaction) for the K2lQ enzyme. The K, for NADP+ was unchanged in both mutant enzymes. The K, for NAD+ was increased 1.5-and 3.2-fold, compared to the wild-type enzyme, in the K21R and K21Q enzymes, respectively. For the K21R enzyme the kc,, for the NAD-and NADP-linked reactions was unchanged. The kc,, for the K21Q enzyme was increased in the NAD-linked reaction by 26% and decreased by 30% in the NADP-linked reaction from the values for the wild-type enzyme. The data are consistent with Lys-21 participating in the binding of the phosphate group of the substrate to the enzyme via charge-charge interaction.

show abstract

Section: Methodsmentioning

confidence: 99%

Lysine‐21 of Leuconostoc mesenteroides glucose 6‐phosphate dehydrogenase participates in substrate binding through charge–charge interaction

Lee

Levy

1992

Protein Science

View full text Add to dashboard Cite

show abstract

“…The DNA sequences of the genes in this report are available in the following references: zwf (27), gnd (24), sodA (38), and nfo (30). The GenBank accession numbers for these sequences are M55005 (zwf), K02072 (gnd), M94879 (sodA), and M22591 (nfo).…”

Section: Methodsmentioning

confidence: 99%

“…Strain NF3079 [dam-3 araD139 ⌬(araABOIC-leu)7679 rpsL galU galK ⌬(lac)X74] was used for the preparation of plasmid DNA requiring restriction with BclI. The properties of strain HB301( DR101), which carries a zwf-lac protein fusion, have been described elsewhere (27).…”

Section: Methodsmentioning

confidence: 99%

“…Plasmid pMLB1034, a derivative of pBR322, has an EcoRI-SmaIBamHI cloning site that can be used for construction of protein fusions to lacZ (34). Plasmids pDR17 (zwf), pCMP78(ϩ) (gnd), and pSOD-1 (sodA) have been described elsewhere (5,27,38). Plasmid pIT3 (nfo) carries the BamHI-EcoRI fragment from pRPC124 (30) in pRS551 (36).…”

Section: Methodsmentioning

confidence: 99%

“…In vivo recombination between plasmids carrying the various protein fusions and RZ5 was used to transfer the fusions to bacteriophage (27). Recombinant bacteriophage were selected as Lac ϩ plaques on strain HB301 plated with top agar on H plates (23) with X-Gal and purified to homogeneity.…”

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Genetic definition of the Escherichia coli zwf "soxbox," the DNA binding site for SoxS-mediated induction of glucose 6-phosphate dehydrogenase in response to superoxide

1995

Self Cite

View full text Add to dashboard Cite

In Escherichia coli K-12, transcription of zwf, the gene for glucose 6-phosphate dehydrogenase, is subject to growth rate-dependent regulation and is activated by SoxS in response to superoxide stress. To define genetically the site of SoxS activation, we undertook a detailed deletion analysis of the zwf promoter region. Using specifically targeted 5 and 3 deletions of zwf sequences, we localized the SoxS activation site to a 21-bp region upstream of the zwf promoter. This minimal ''soxbox'' was able to confer paraquat inducibility when placed upstream of a normally unresponsive gnd-lacZ protein fusion. In addition, we used these findings as the basis for resecting unnecessary sequences from the region upstream of the promoters of two other SoxSregulated genes, sodA and nfo. Like the zwf soxbox, the regions required for SoxS activation of sodA and nfo appear to lie just upstream of or overlap the ؊35 hexamers of the corresponding promoters. Importantly, the sequence boundaries established here by deletion analysis agree with the primary SoxS recognition sites of zwf, sodA, and nfo that we previously identified in vitro by gel mobility shift and DNase I protection assays with a purified MalE-SoxS fusion protein.In Escherichia coli, the oxidative branch of the pentose phosphate pathway provides ribose for nucleoside biosynthesis and NADPH for reductive biosynthesis (9, 12). Two of the enzymes of this pathway, glucose 6-phosphate dehydrogenase (encoded by zwf) and 6-phosphogluconate dehydrogenase (encoded by gnd) are subject to growth rate-dependent regulation (40). The specific activities of these enzymes increase in proportion to increased growth rate during steady-state growth on different carbon sources. Studies on the growth rate-dependent regulation of zwf and gnd have shown that at least part of the increase in the specific activities of these enzymes is attributable to an increase in the transcription rates of the respective genes (25,26).Apart from its basic growth rate-dependent regulation, zwf is also a member of at least two other regulons. Thus, unlike gnd, zwf expression is transcriptionally activated by SoxS during episodes of oxidative stress induced by exposure of E. coli to superoxide-generating agents such as paraquat (17,18,39). Indeed, E. coli strains deleted for zwf are hypersensitive to oxidative stress, thus making zwf a pivotal member of the soxRS regulon (18, 21). In addition, treatment of cells with several antibiotics and aromatic weak acids appears to activate zwf transcription through the action of marA, thus making zwf a member of the multiple antibiotic resistance (mar) regulon (2,6,7,14,16).The work presented here has several overlapping long-range objectives. One is to understand the physiological basis for zwf's membership in the soxRS and marRAB regulons. The other is to identify the cis-acting regulatory sites of zwf that are involved in superoxide control for subsequent comparison with the sites required for growth rate control and for induction by antibiotics and aromatic weak...

show abstract

Co‐production of hydrogen and ethanol from glucose by modification of glycolytic pathways in Escherichia coli – from Embden‐Meyerhof‐Parnas pathway to pentose phosphate pathway

Seol

Sekar

Raj

et al. 2016

Biotechnology Journal

View full text Add to dashboard Cite

Hydrogen (H2) production from glucose by dark fermentation suffers from the low yield. As a solution to this problem, co-production of H2 and ethanol, both of which are good biofuels, has been suggested. To this end, using Escherichia coli, activation of pentose phosphate (PP) pathway, which can generate more NADPH than the Embden-Meyhof-Parnas (EMP) pathway, was attempted. Overexpression of two key enzymes in the branch nodes of the glycolytic pathway, Zwf and Gnd, significantly improved the co-production of H2 and ethanol with concomitant reduction of pyruvate secretion. Gene expression analysis and metabolic flux analysis (MFA) showed that, upon overexpression of Zwf and Gnd, glucose assimilation through the PP pathway, compared with that of the EMP or Entner-Doudoroff (ED) pathway, was greatly enhanced. The maximum co-production yields were 1.32 mol H2 mol(-1) glucose and 1.38 mol ethanol mol(-1) glucose, respectively. It is noteworthy that the glycolysis and the amount of NAD(P)H formed under anaerobic conditions could be altered by modifying (the activity of) several key enzymes. Our strategy could be applied for the development of industrial strains for biological production of reduced chemicals and biofuels which suffers from lack of reduced co-factors.

show abstract

Molecular characterization of the Escherichia coli K-12 zwf gene encoding glucose 6-phosphate dehydrogenase

Cited by 90 publications

References 57 publications

Lysine‐21 of Leuconostoc mesenteroides glucose 6‐phosphate dehydrogenase participates in substrate binding through charge–charge interaction

Lysine‐21 of Leuconostoc mesenteroides glucose 6‐phosphate dehydrogenase participates in substrate binding through charge–charge interaction

Genetic definition of the Escherichia coli zwf "soxbox," the DNA binding site for SoxS-mediated induction of glucose 6-phosphate dehydrogenase in response to superoxide

Co‐production of hydrogen and ethanol from glucose by modification of glycolytic pathways in Escherichia coli – from Embden‐Meyerhof‐Parnas pathway to pentose phosphate pathway

Contact Info

Product

Resources

About