STRUCTURE AND FUNCTION OF CARBAMOYLPHOSPHATE SYNTHETASE - ON MECHANISM OF BICARBONATE ACTIVATION

The Km‐values for ammonium and bicarbonate in carbamoylphosphate synthesis are 1.1 mM and 5.3 mM respectively at saturation with ATP and acetylglutamate. The are independent of the concentration of the second substrate. The same Km for bicarbonate is found in the hydrolysis of ATP, which is catalyzed by the enzyme in the absence of ammonium. The [32P]orthophosphate–ATP‐exchange, which is very slow, requires bicarbonate and ammonium. With these findings, the following mechanisms for the C–N‐ligase step(s) of the enzyme are possible: (a) enzyme‐bound carbamate is formed by a simultaneous reaction of ATP, bicarbonate and ammonium; (b) at first enzyme‐bound “active CO2” is formed in a rapid and reversible reaction, which releases both the ATP‐products, ADP and orthophosphate only upon the reaction with ammonium. Possible structures for this intermediate would be carboxyphosphate plus ADP and the addition compound of bicarbonate with ATP (with penta‐coordinated phosphorus). The participation of biotin was excluded. As can be estimated from the kinetic findings, only a small fraction of the enzyme molecules would ever be charged with “activity, CO2”; the half‐time of its decomposition, as calculated from the ATPase activity, would be ≤ 2 sec. Copyright © 1969, Wiley Blackwell. All rights reserved