MOUSE NEUROBLASTOMA CELL ADENYLATE-CYCLASE - REGULATION BY 2-CHLOROADENOSINE, PROSTAGLANDIN-E1 AND CATIONS MG2+, CA2+ AND MN2+

Some basic kinetic properties of adenylate cyclase in cell free preparations of mouse neuroblastoma were investigated. Production of c [cyclic] AMP from ATP by the enzyme required the presence of either Mg2+ or Mn2+ in addition to ATP. In the presence of Mg2+ the Km for ATP was 120 .+-. 15 .mu.M and the interaction of ATP and adenylate cyclase appeared to be non-cooperative (Hill coefficient of 1). Mg2+ concentrations in excess of the ATP concentration caused stimulation, although similar excess concentrations of Mn2+ cause inhibition. Prostaglandin E1 and 2-chloroadenosine activated the enzyme. The Km of the cyclase for 2-chloroadenosine was 6 .mu.M. Activation by 2-chloroadenosine led to an increase in Vmax but did not affect the Km for ATP. At a fixed ATP concentration the extent of activation caused by prostaglandin E1 and 2-chloroadenosine was inversely related to the Mg2+ concentration. Ca2+ caused inhibition of adenylate cyclase from 0.1-4 mM with a Ki of 5 .times. 10-4 M. Ca2+ interaction with the enzyme in the absence or presence of either 2-chloroadenosine or prostaglandin E1 appeared cooperative (i.e., Hill coefficients of .apprx. 2). Ca2+ inhibition was non-competitive with respect to either ATP or 2-chloroadenosine but was progressively diminished by increasing Mn2+ concentrations. Divalent cation effects and activation by 2-chloroadenosine and prostaglandin E1 of the neuroblastoma adenylate cyclase were compared with ion effects and hormone activation of the enzyme obtained from non-neuronal tissue.