Enzymes in facultative anaerobiosis of molluscs-II. Basic catalytic properties of phosphoenolpyruvate carboxykinase in oyster adductor muscle

Abstract

1. 1. p-Enolpyruvate carboxykinase (E.C. 4.1.1.32) from oyster adductor tissue occurs as a single activity peak with a pI value of 6·64. 2. 2. The enzyme is present in high activities only in adductor tissue. 3. 3. It exhibits a relatively high degree of specificity for IDP and GDP and is inactive in the presence of IMP, GMP and ADP. Oyster enzyme, unlike p-enolpyruvate carboxykinases from other sources, exhibits no activity in the presence of Mg 2+. 4. 4. The pH activity profiles in the presence of Zn 2+ and Mn 2+ are similar in general pattern but the pH optima are 5·1 and 6·0 respectively. 5. 5. Kinetic constants for metal ions, IDP and p-enolpyruvate were determined at different pH values. For metal ions, the apparent enzyme-Mn 2+ affinity is greater at pH 6·0 than at pH 5·1 while the apparent enzyme-Zn 2+ affinity is greater at pH 5·1 than at pH 6·0. S 0·5 values for both metal ions under different conditions vary between 0·15 and 0·52 mM. The absolute values of S 0·5 for IDP are comparable in the presence of Zn 2+ and Mn 2+ (in the range of 0·03–0.06 mM) and these are largely independent of pH. p-Enolpyruvate saturation exhibits aberrant behaviour in the present of Mn 2+ while in the presence of Zn 2+ it follows Michaelis-Menten patterns. Equally significant, the apparent K m values are at least one order of magnitude less than observed for the enzyme in the presence of Mn 2+. 6. 6. Low concentrations of Cu 2+ (10–40 μM) inhibit the enzyme in the presence of either Zn 2+ or Mn 2+. Cu 2+ in the presence of Mn 2+ exhibits a competitive inhibition pattern while in the presence of Zn 2+, a non-competitive inhibition pattern occurs. 7. 7. The physiological role of the enzyme is discussed in relation to its cellular distribution, metal ion requirements and pH properties.