PMID:26327
| Citation |
Sullivan, JJ, Muchnicky, EE, Davidson, BE and Jago, GR (1977) Purification and properties of the pyrrolidonecarboxylate peptidase of Streptococcus faecium. Aust. J. Biol. Sci. 30:543-52 |
|---|---|
| Abstract |
Pyrrolidonecarboxylate peptidase (EC 3.4.11.8) from Streptococcus faecium was purified by fractionation with streptomycin sulphate and ammonium sulphate, by chromatography on Sephadex G200 and DEAE-cellulose, and by preparative electrophoresis on Sephadex G25. The purified enzyme on acrylamide gel showed a strong protein band which contained enzyme activity and a very faint band which had no activity. The subunit molecular weight of the purified enzyme was estimated by acrylamide gel electrophoresis in sodium dodecyl sulphate to be 42,000 +/- 1,000. The enzyme showed optimum activity at pH 7.6 and was unstable in the absence of 2-mercaptoethanol. The sensitivity of the enzyme to alkylating agents (N-ethylmaleimide and iodoacetamide) suggested that free sulphydryl groups were essential for enzyme activity. The enzyme was rapidly inactivated above 45 degrees C. The values of the Michaelis constants (Km) obtained with various L-pyrrolidonecarboxylyl dipeptides were similar although there was a 10-fold range in the maximal rates of hydrolysis of these substrates. Inhibition studies showed that the substrate analogues 2-pyrrolidone and pyrrolidonecarboxylate are competitive inhibitors of the enzyme. The binding of substrates and inhibitors to the active site of the enzyme is discussed. |
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| Keywords |
Aminopeptidases/isolation & purification; Drug Stability; Hydrogen-Ion Concentration; Kinetics; Molecular Weight; Pyroglutamyl-Peptidase I/antagonists & inhibitors; Pyroglutamyl-Peptidase I/isolation & purification; Streptococcus/enzymology; Substrate Specificity |
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