Topic
Methylglyoxal reductase
About: Methylglyoxal reductase is a research topic. Over the lifetime, 32 publications have been published within this topic receiving 740 citations.
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TL;DR: Three different enzymes were genetically fused with streptavidin binding peptide, Spy and Halo-based tags, to enable their specific and directional immobilization on magnetic microbeads coated with complementary receptors for modular configuration of microfluidic packed-bed reactors.
Abstract: The establishment of microfluidic enzyme cascades is a topical field of research and development, which is currently hampered by the lack of methodologies for mild and efficient immobilization of isolated enzymes. We here describe the use of self-immobilizing fusion enzymes for the modular configuration of microfluidic packed-bed reactors. Specifically, three different enzymes, the (R)-selective alcohol dehydrogenase LbADH, the (S)-selective methylglyoxal reductase Gre2p and the NADP(H) regeneration enzyme glucose 1-dehydrogenase GDH, were genetically fused with streptavidin binding peptide, Spy and Halo-based tags, to enable their specific and directional immobilization on magnetic microbeads coated with complementary receptors. The enzyme-modified beads were loaded in four-channel microfluidic chips to create compartments that have the capability for either (R)- or (S)-selective reduction of the prochiral CS-symmetrical substrate 5-nitrononane-2,8-dione (NDK). Analysis of the isomeric hydroxyketone and ...
78 citations
TL;DR: It is proposed that the predominant mechanism for methylglyoxal detoxification in the African trypanosome is via the methyl Glyoxal reductase pathway to l‐lactate.
Abstract: The glyoxalase system, comprising the metalloenzymes glyoxalase I (GLO1) and glyoxalase II (GLO2), is an almost universal metabolic pathway involved in the detoxification of the glycolytic byproduct methylglyoxal to d-lactate. In contrast to the situation with the trypanosomatid parasites Leishmania major and Trypanosoma cruzi, this trypanothione-dependent pathway is less well understood in the African trypanosome, Trypanosoma brucei. Although this organism possesses a functional GLO2, no apparent GLO1 gene could be identified in the T. brucei genome. The absence of GLO1 in T. brucei was confirmed by the lack of GLO1 activity in whole cell extracts, failure to detect a GLO1-like protein on immunoblots of cell lysates, and lack of d-lactate formation from methylglyoxal as compared to L. major and T. cruzi. T. brucei procyclics were found to be 2.4-fold and 5.7-fold more sensitive to methylglyoxal toxicity than T. cruzi and L. major, respectively. T. brucei also proved to be the least adept of the 'Tritryp' parasites in metabolizing methylglyoxal, producing l-lactate rather than d-lactate. Restoration of a functional glyoxalase system by expression of T. cruzi GLO1 in T. brucei resulted in increased resistance to methylglyoxal and increased conversion of methylglyoxal to d-lactate, demonstrating that GLO2 is functional in vivo. Procyclic forms of T. brucei possess NADPH-dependent methylglyoxal reductase and NAD(+)-dependent l-lactaldehyde dehydrogenase activities sufficient to account for all of the methylglyoxal metabolized by these cells. We propose that the predominant mechanism for methylglyoxal detoxification in the African trypanosome is via the methylglyoxal reductase pathway to l-lactate.
74 citations
TL;DR: Two new products were formed in addition to ethanol, acetate, H2 and CO2: d (-)-1,2-propanediol and d(-)-lactate, apparently synthesized via the methylglyoxal by-pass in C. sphenoides grown on glucose in a phosphate-limited medium.
Abstract: Clostridium sphenoides was grown on glucose in a phosphate-limited medium. Below 80 μM phosphate two new products were formed in addition to ethanol, acetate, H2 and CO2: d(-)-1,2-propanediol and d(-)-lactate. These compounds were apparently synthesized via the methylglyoxal by-pass. The activity of the enzymes involvedmethylglyoxal synthase, methylglyoxal reductase, 1,2-propanediol dehydrogenase and glyoxalase-could be demonstrated in cell extracts of C. sphenoides. The formation of 1,2-propanediol from methylglyoxal proceeded via lactaldehyde. The enzyme methylgloxal synthase was inhibited by phosphate.
67 citations
TL;DR: An enzyme catalyzing the reduction of methylglyoxal was isolated from Saccharomyces cerevisiae and its enzymatic properties were analyzed and it was tentatively designated the enzyme methyl glyoxal reductase.
Abstract: An enzyme catalyzing the reduction of methylglyoxal was isolated from Saccharomyces cerevisiae and its enzymatic properties were analyzed. The enzyme, specifically eluted from a blue-dextran – Sepharose CL-6B column by the substrate, methyglyoxal, was homogeneous on polyacrylamide gel electrophoresis. The enzyme consisted of single polypeptide chain with a relative molecular mass of 43 000. The enzyme was glycoprotein and contained 6.6% carbohydrate. NADPH was specifically required for activity and the Km for NADPH was 2.0 × 10−7 M. The enzyme was active on various glyoxals such as glyoxal, methylglyoxal (Km= 5.88 mM) and phenylglyoxal (Km= 1.54 mM). The reaction catalyzed by the enzyme was virtually irreversible. The activity was inhibited by sulfhydryl agents and activated by reducing agents such as glutathione. Intermediates in glycolysis, nucleosides, nucleotides, polyamines and various metal ions showed little inhibitory or activating effects on enzyme activity. Tricarboxylic acids showed a slight inhibitory effect. The activity of the enzyme was strongly inhibited by anionic detergents. The enzyme was rapidly inactivated by incubating with the substrates probably because of the non-enzymatic interaction between glyoxals and NH2 groups in arginine residues in the enzyme. NADP, one of the reaction products, also inhibited the enzyme activity and the Ki for NADP was about 0.07 mM. We tentatively designated the enzyme methylglyoxal reductase.
63 citations
TL;DR: It is shown that methylglyoxal reductase (NADPH‐dependent) is encoded by GRE2 (YOL151w) and associated this activity with its gene by partially purifying the enzyme and identifying by MALDI–TOF the proteins in candidate bands on SDS–PAGE gels whose relative intensities correlated with specific activity through three purification steps.
Abstract: Methylglyoxal is associated with a broad spectrum of biological effects, including cytostatic and cytotoxic activities. It is detoxified by the glyoxylase system or by its reduction to lactaldehyde by methylglyoxal reductase. We show that methylglyoxal reductase (NADPH-dependent) is encoded by GRE2 (YOL151w). We associated this activity with its gene by partially purifying the enzyme and identifying by MALDI-TOF the proteins in candidate bands on SDS-PAGE gels whose relative intensities correlated with specific activity through three purification steps. The candidate proteins were then purified using a glutathione-S-transferase tag that was fused to them, and tested for methylglyoxal reductase activity. The advantage of this approach is that only modest protein purification is required. Our approach should be useful for identifying many of the genes that encode the metabolic pathway enzymes that have not been associated with a gene (about 275 in S. cerevisiae, by our estimate).
61 citations