PURIFICATION AND CHARACTERIZATION OF PROTEIN PHOSPHATASE 2C IN RAT PAROTID ACINAR-CELLS - 2 FORMS OF MG2-ACTIVATED HISTONE PHOSPHATASE AND PHOSPHORYLATION BY CAMP-DEPENDENT PROTEIN-KINASE()

Two forms of Mg2+-activated histone phosphatase activities were partia lly purified from rat parotid acinar cells using Mono Q and gel filtra tion chromatography. Both enzymes activities were dependent on the pre sence of Mg2+, showing little activity in the presence of EDTA. The ac tivities fractionated on the Mono Q column into two peaks: the first w as a minor peak of histone phosphatase activity; the second was a majo r peak. These two peaks eluted at distinct positions on the gel filtra tion column. The molecular masses of the two peak fractions correspond ed to 46 and 55 kDa, respectively on SDS-gels. The first 46-kDa peak i mmunoreacted with anti-PP2C alpha phosphatase antibody and like PP2C a lpha phosphatase could be phosphorylated by cAMP-dependent protein kin ase. The second 55-kDa peak showed neither reactivity with anti-PP2C a lpha phosphatase antibody nor phosphorylability by cAMP-dependent prot ein kinase, but retained a Mg2+ or Mn2+ dependance for its histone pho sphatase activity. Ca2+ showed a strong inhibition on this activity. O n the basis of these observations, we have identified the first peak e nzyme as PP2C alpha phosphatase and the second peak as a novel PP2C-li ke phosphatase. (C) 1996 Academic Press Inc.