Histidyl phosphorylation and dephosphorylation of P36 in rat liver extract.
Protein histidine kinase (Motojima, K., and Goto, S. (1993) FEBS Lett. 319, 75-79) and phosphatase in rat liver extract were characterized. The histidine kinase was recovered mostly in the membrane and the phosphatase in the soluble fraction. The kinase and its substrate 36-kDa protein (P36) were co-solubilized from the membrane under conditions in which most of the other kinases, and their substrate proteins were not solubilized. The solubilized kinase and P36 were co-eluted after high pressure liquid chromatography gel filtration, showing an apparent molecular mass of 70-75 kDa. They were also co-eluted after ion exchange chromatography. These characteristics, together with its complete resistance to genistein, indicate that the rat liver histidine kinase is not cognate to the yeast enzyme (Huang, J., Nasr, M., Kim, Y., and Matthews, H.R. (1992) J. Biol. Chem. 267, 15511-15515). The phosphatase that dephosphorylates histidyl-phosphorylated P36 was also studied using rat liver subcellular fractions and in vitro phosphorylated P36 as the substrate. The characteristics of the phosphatase, that is, 1) Mg2+ requirement for activity, 2) apparent molecular mass of 45 kDa by high performance liquid chromatography gel filtration, and 3) resistance to 100 microM okadaic acid, suggest that the primary phosphatase active in vitro is protein phosphatase 2C.
Motojima
K
K
Department of Biochemistry, School of Pharmaceutical Sciences, Toho University, Chiba, Japan.
Goto
S
S
D000818:Animals; D002994:Clofibrate / Q000494:pharmacology; D019833:Genistein; D006639:Histidine / Q000031:analogs & derivatives* / Q000378:metabolism; D000071677:Histidine Kinase; D007529:Isoflavones / Q000494:pharmacology; D008099:Liver / Q000201:enzymology*; D008297:Male; D008970:Molecular Weight; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D011916:Rats, Inbred F344; D013347:Subcellular Fractions / Q000378:metabolism; D013997:Time Factors
D007529:Isoflavones; D010750:Phosphoproteins; D006639:Histidine; D019833:Genistein; D011494:Protein Kinases; D000071677:Histidine Kinase; D010749:Phosphoprotein Phosphatases; D002994:Clofibrate; C005436:phosphohistidine