Reinvestigation of the role of thiol groups of glyoxalase I purified from yeast (Saccharomyces cerevisiae).
Glyoxalase I has been purified to homogeneity from Saccharomyces cerevisiae and tested with two different thiol reagents, i.e., 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) and 1-chloro-2,4-dinitrobenzene (CDNB). DTNB reacts with four thiol groups per molecule of enzyme and leads to a complete inhibition which is not reversed by addition of the disulfide-reducing agent dithiothreitol. On the other hand, CDNB slightly affects the glyoxalase-I activity and alkylates only one thiol residue/enzyme. In agreement, DTNB reacts with three thiol residues of the CDNB-reacted enzyme and no reactivation is observed after dithiothreitol treatment. The peptide containing the CDNB-reactive thiol group has been isolated and the sequence overlaps the segment 58-63 of the only known primary structure of glyoxalase I from Pseudomonas putida.
Dipartimento di Biologia, Università di Roma Tor Vergata, Italy.
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Caccuri
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Risitano
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Marmocchi
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Venardi
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Aureli
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Petruzzelli
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Desideri
D000595:Amino Acid Sequence; D004137:Dinitrochlorobenzene; D004228:Dithionitrobenzoic Acid; D004229:Dithiothreitol; D007791:Lactoylglutathione Lyase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013438:Sulfhydryl Compounds / Q000737:chemistry*; D014357:Trypsin
D004137:Dinitrochlorobenzene; D013438:Sulfhydryl Compounds; D004228:Dithionitrobenzoic Acid; D014357:Trypsin; D007791:Lactoylglutathione Lyase; D004229:Dithiothreitol