Effect of chemical modification of sulfhydryl groups of human erythrocyte enzymes.
Erythrocyte lysate proteins by five methods were chemically modified by methanethiolation with methyl methanethiolsulfonate (MMTS). This relatively newly studied agent delivers the small uncharged, non-hydrogen-bonding CH3S-group to accessible -SH groups under mild conditions. Glycolytic and nonglycolytic red cell enzymes studied could be grouped into four categories based on degree of catalytic inhibition by MMTS, which ranged from 100% to nil. NADH ((0.06 mM) protected against lactate dehydrogenase (LDH) inhibition nearly completely, and partial protection was evident at 1 microM concentrations. NAD was virtually ineffective at comparable molarities, and pyruvate failed to protect significantly. Glucose-6-phosphate dehydrogenase (G-6-PD) (but not 6-PGD) was protected by NADP (but minimally if at all by NADPH) at low molarities. Substrate G-6-P failed to protect. Mg++ alone protected enolase to a major degree against MMTS inhibition. Residual activities of several enzymes were rendered highly unstable to thermal stress, and in the case of selected enzymes, alteration of pH activity curves and catalytic activity at diminished substrate concentration was demonstrable. The highly specific modification of sulfhydryl groups of cysteinyl residues under mild conditions preserving other aspects of enzyme structure provides insights into the role of these most reactive of all amino side-chains in the enzyme proteins derived from human red cell lysates.
D002384:Catalysis; D004912:Erythrocytes / Q000201:enzymology*; D005954:Glucosephosphate Dehydrogenase / Q000037:antagonists & inhibitors / Q000097:blood; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007770:L-Lactate Dehydrogenase / Q000037:antagonists & inhibitors / Q000097:blood; D008741:Methyl Methanesulfonate / Q000031:analogs & derivatives* / Q000494:pharmacology; D009249:NADP / Q000494:pharmacology; D010751:Phosphopyruvate Hydratase / Q000037:antagonists & inhibitors / Q000097:blood; D013438:Sulfhydryl Compounds / Q000494:pharmacology
D013438:Sulfhydryl Compounds; C014674:methyl methanethiosulfonate; D009249:NADP; D008741:Methyl Methanesulfonate; D007770:L-Lactate Dehydrogenase; D005954:Glucosephosphate Dehydrogenase; D010751:Phosphopyruvate Hydratase