Excess zinc ions are a competitive inhibitor for carboxypeptidase A

Abstract

The mechanism for inhibition of enzyme activity by excess zinc ions has been studied by kinetic and equilibrium dialysis methods at pH 8.2, I = 0.5 M. With carboxypeptidase A (bovine pancreas), peptide (carbobenzoxyglycyl-L-phenylalanine and hippuryl-L-phenylalanine) and ester (hippuryl-L-phenyl lactate) substrates were inhibited competitively by excess zinc ions. The Ki values for excess zinc ions with carboxypeptidase A at pH 8.2 are all similar [Ki = (5.2-2.6) X 10(-5) M]. The apparent constant for dissociation of excess zinc ions from carboxypeptidase A was also obtained by equilibrium dialysis at pH 8.2 and was 2.4 X 10(-5) M, very close to the Ki values above. With arsanilazotyrosine-248 carboxypeptidase A ([(Azo-CPD)Zn]), hippuryl-L-phenylalanine, carbobenzoxyglycyl-L-phenylalanine, and hippuryl-L-phenyl lactate were also inhibited with a competitive pattern by excess zinc ions, and the Ki values were (3.0-3.5) X 10(-5) M. The apparent constant for dissociation of excess zinc ions from arsanilazotyrosine-248 carboxypeptidase A, which was obtained from absorption changes at 510 nm, was 3.2 X 10(-5) M and is similar to the Ki values for [(Azo-CPD)Zn]. The apparent dissociation and inhibition constants, which were obtained by inhibition of enzyme activity and spectrophotometric and equilibrium dialysis methods with native carboxypeptidase A and arsanilazotyrosine-248 carboxypeptidase A, were almost the same. This agreement between the apparent dissociation and inhibition constants indicates that the zinc binding to the enzymes directly relates to the inhibition of enzyme activity by excess zinc ions. Excess zinc ions were competitive inhibitors for both peptide and ester substrates.(ABSTRACT TRUNCATED AT 250 WORDS)

A structural and functional analysis of Nna1 inPurkinje cell degeneration(pcd) mice

“…7C). This biphasic property is observed in other Zn 2ϩ -dependent enzymes (34,35). Zinc carboxypeptidases are inhibited by divalent cation chelators (36,37), and both EDTA and OP, inhibitors of zinc carboxypeptidases (37,38), reduce the activity of Nna1 (Fig.…”

Rat kidney endopeptidase 24.16

“…The fact that endopeptidase 24.16 was blocked by zinc while the most potent inhibitor agent was a-phenanthroline suggested that renal endopeptidase 24.16 belongs to the zinc-metallopeptidase family, in agreement with our previous data showing that the intestinal apoenzyme could be eficiently reactivated by low concentrations of zinc (Barelli et al, 1988). The dual activating and inactivating effect of zinc ions was previously reported for various zinc-containing metalloenzymes and was recently examined by Hirose et al (1987) who showed that excess zinc ions could act as competitive inhibitor of carboxypeptidase A.…”

The ς 70 Transcription Factor TyrR Has Zinc-Stimulated Phosphatase Activity That Is Inhibited by ATP and Tyrosine

“…First, zinc ions are required for the enzymatic activities of both proteins; second, excess Zn 2ϩ inhibits both the enzymatic activity of carboxypeptidase and the phosphatase activity of TyrR (25); third, upon adding zinc ion to metal-free apoenzymes, both proteins showed a decrease in fluorescence intensity and a slight change in emission wavelength (14) (Fig. 8); and fourth, both proteins lack cysteine-or histidine-rich regions as identifiable zinc-binding motifs.…”