[Demonstration, kinetic study and solubilization of particular adenylate cyclase from Nocardia restricta]

An adenylate cyclase (EC r.6.1.1.) was found in cell-free extracts of several Nocardia species. The enzyme from Nocardia restricta has been specially studied. It is a membrane enzyme which exhibits a strong specific activity, one hundred times greater than that of mammals. It has an optimal pH of 8.5 in Tris buffer and an absolute requirement for divalent ions, Mg++ or Mn++ (Mn++ ions are the most efficient). The kinetic properties of this adenylate cyclase are similar to those that could be expected of an allosteric enzyme having, as a substrate, the ATP-Mg++ complex and, as an activator, free Mn++ ions. Ca++ ions are activators: they set up the maximum velocity without modification of the KM. GTP is a competitive inhibitor (KI = 5.10(-5) M). Fluoride ions have no detectable effect on activity. Non-ionic detergents, Lubrol WX and Triton X 100, are inhibitors of the enzyme which has been partially solubilized by repeated freezing and thawing, following by brief ultra-sonic treatment. Catalytic sites are not modified after the solubilization, but cooperative effects between moles of substrate ATP-Mn++ are diminished: the KM becomes smaller and the sigmoidal shape of the curve v = f (ATP-Mn++) is attenuated.