The identification of a structurally important cysteine residue in the glycerol dehydrogenase from Bacillus stearothermophilus.
Evidence is presented to demonstrate that the Zn2+ metallo-enzyme glycerol dehydrogenase from the thermophile Bacillus stearothermophilus has one cysteine residue per subunit which is only available for reaction with thiol reagents in the metal-depleted form of the enzyme. Modification of the metal-depleted enzyme by methyl methanethiosulphonate prevents the reactivation of the enzyme by Zn2+ ions and induces dissociation of the oligomer into subunits. The rate of reaction of the cysteine residue with the thiol reagent DTNB is limited by a factor other than reagent concentration and it is proposed that the reagent only reacts with the cysteine residue in dissociated monomers. The enzyme has been labelled at the single cysteine residue by radioactive iodo[2-3H]acetic acid. Two radiolabelled peptides have been isolated and sequenced; one peptide is a component of the other. Spectroscopic evidence suggests that the cysteine residue is not involved in ligation of the essential metal ion. Chemical modification studies using the reagent diethylpyrocarbonate have suggested that two histidines are involved in the ligation of the metal.
Spencer
P
P
Department of Biochemistry, University of Southampton, U.K.
Scawen
M D
MD
Atkinson
T
T
Gore
M G
MG
D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid; D003035:Cobalt / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry*; D004047:Diethyl Pyrocarbonate; D004228:Dithionitrobenzoic Acid; D001411:Geobacillus stearothermophilus / Q000201:enzymology*; D006639:Histidine; D007461:Iodoacetates; D019807:Iodoacetic Acid; D007700:Kinetics; D046911:Macromolecular Substances; D008741:Methyl Methanesulfonate / Q000031:analogs & derivatives / Q000494:pharmacology; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D013401:Sugar Alcohol Dehydrogenases / Q000737:chemistry* / Q000378:metabolism; D014316:Tritium; D014357:Trypsin; D015032:Zinc / Q000494:pharmacology
D007461:Iodoacetates; D046911:Macromolecular Substances; D010446:Peptide Fragments; D014316:Tritium; C014674:methyl methanethiosulfonate; D003035:Cobalt; D006639:Histidine; D004228:Dithionitrobenzoic Acid; D008741:Methyl Methanesulfonate; D013401:Sugar Alcohol Dehydrogenases; C020672:glycerol dehydrogenase; D014357:Trypsin; D015032:Zinc; D003545:Cysteine; D004047:Diethyl Pyrocarbonate; D019807:Iodoacetic Acid