Kinetics of irreversible enzyme inhibition by an unstable inhibitor

Abstract

A mathematical treatment for the general case of enzyme inactivation by an inhibitor that breaks down in solution in a first-order reaction is presented. Cathepsin D was inactivated by fluorescein isothiocyanate with a K(i) of 4.47mum. Kinetic constants were also determined for the inactivation of cathepsin D by 1,1-bis(diazoacetyl)-2-phenylethane, and the inactivation of pepsin C by diazoacetyl-dl-norleucine methyl ester.

Kinetic analysis of the inhibition of sulfate transport in human red blood cells by isothiocyanates

“…alternative mechanism for the inactivation process would be the covalent attachment of the inhibitor on the membrane sites, without the prior formation of a reversible membrane site-inhibitor complex, that is, by a simple bimolecular mechanism (Baker, 1967;Rakitzis, 1974 (Cabantchik & Rothstein, 1972, 1974Lepke et al, 1976;Zaki et al, 1975). The reversible effect of isothiocyanates on the flux constant (kr) appears to be quite a weak one in the case of INDS (Table 1).…”

“…These values are approximately 20~o smaller than the corresponding values when the isothiocyanate was dissolved in the sulfate-chloride medium immediately before the beginning of the flux determination (Tables 1 and 2). The isothiocyanates will hydrolyze to the corresponding amino-arylsulfonates which are not known to bind covalently to the red cell membranes, but they are known to act as reversible inhibitors (Cabantchik & Rothstein, 1972, 1974Zaki et al, 1975).…”

Activation of cathepsin D by glycine ethyl ester

“…The final pH of the incubation mixture was determined with a Radiometer 29 pH-meter. The rest of the assay procedure for cathepsin D activity was as described previously (Rakitzis, 1974).…”

Kinetics of protein modification reactions

“…The assumption is made that the modifying agent disappears in a first-order reaction, so that (16)-(19) has been developed by Purdie & Heggie (1969), and by Ashani et al (1972). This treatment has been applied to cases of enzyme inactivation by unstable modifying agents by Ashani et al (1972), Maglothin & Wilson (1974), Rakitzis (1974), Maglothin et al (1975), Barnett & Rosenberry (1978), Johnson & Poisner (1980), and by Makoff & Malcolm (1980.…”