Activation of snake venom metalloproteinases by a cysteine switch-like mechanism

• Published in: FEBS letters 1993-11, Vol.335 (1), p.76-80
• Main Authors: Grams, Frank, Huber, Robert, Kress, Lawrence F., Moroder, Luis, Bode, Wolfram
• Format: Article
• Language: English
• Subjects: Abz, 2-aminobenzoyl; Adamalysin; Amino Acid Sequence; Analytical, structural and metabolic biochemistry; Binding Sites; Biological and medical sciences; Conserved Sequence; Crotalid Venoms - metabolism; Crotalus adamanteus; Cysteine - metabolism; Cysteine switch; Enzyme Activation; Enzymes and enzyme inhibitors; FAB-MS, fast atom bombardment mass spectroscopy; Fundamental and applied biological sciences. Psychology; Glutathione - pharmacology; Hydrogen-Ion Concentration; Hydrolases; Matrix metalloproteinase; Metalloendopeptidases - antagonists & inhibitors; Metalloendopeptidases - chemistry; Metalloendopeptidases - metabolism; Molecular Sequence Data; Peptide Fragments - chemistry; Peptide Fragments - pharmacology; Protein Structure, Secondary; Proteinase activation; Snake venom metalloproteinase; Stability of cysteine peptide; standard abbreviations for amino acids and peptide derivatives are used as recommended by the IUPAC-IUB commission on biochemical nomenclature. The amino acids are of l-configuration unless stated otherwise. HPLC, high-pressure liquid chromatography; Zinc - metabolism
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/0014-5793(93)80443-X

The cDNAs of several snake venom zinc endopeptidases code for a putative propeptide, which includes the conserved cysteine-containing sequence PKMCGVT. It has been suggested that binding of the cysteine thiol function to the active-site zinc, resulting in inactivation of the catalytic domain, occurs in a mode similar to the ‘cysteine switch’ mechanism proposed for matrix metalloproteinases. In order to confirm this hypothesis, inhibition kinetics have been performed on the metalloproteinase adamalysin II of the venom of the snake Crotalus adamanteus using several cysteine peptides. Among these the synthetic hexapeptide PKMCGV-NH 2, corresponding to the conserved sequence portion of the known propeptides, was found to be by far the strongest inhibitor of this proteinase with a K i of 3.4 μM. The inhibitory potencies of an equivalent peptide with the l-Cys replaced by a d-Cys or by an l-Ser as well as of reduced glutathione, cysteine and two unrelated cysteine peptides were by one to two orders of magnitudes lower. These findings strongly support a cysteine switch-like mechanism even for activation of the snake venom metalloproteinases.