Poly(adenylate) polymerase activity of rat brain

Abstract

The poly(adenylate)[poly(A)] polymerase of rat brain, as in rat liver, is located primarily in the nuclear sap when nuclei are prepared under hypertonic conditions. The enzyme can be released from nuclei in two forms. Form I is prepared by gentle incubation of nuclei at 0 degrees C in hypotonic buffer. It has a Mn optimum of 0.6 mM and a pH optimum between 8 and 9. The ATP concentration curve plateaus at 0.2 mM. The optimal poly(A) primer concentration is 600 micrograms/ml, which is three times higher than that for the enzyme similarly prepared from liver. The time course of the reaction for the form I enzyme is increasing over the first 40 min and becomes nearly linear thereafter. Form I is not stimulated by either calcium or cyclic nucleotides, but is inhibited by polyamines, pyrophosphate, and high concentrations of GTP. Form II enzyme is prepared by homogenization of nuclei in hypotonic buffer. It has the same ATP and poly(A) optima as the form I enzyme but displays linear kinetics over a 60-min time course. It is slightly stimulated by cGMP and cAMP and strongly inhibited by spermine, sodium pyrophosphate, and high concentrations of GTP.