Role of protein kinase in cell cycle regulation

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1) In an attempt to elucidate the role of protein kinase C (C kinase) in cell cycle regulation, the direct target of this enzyme was surveyed using the extract of Xenopus laevis oocytes.2) An excellent phosphate acceptor protein with molecular mass of about 25,000 (25kDa protein) was detected in cytosolic fraction of Xenopus oocytes employing C kinase and casein Kinase II as catalyst3) This 25KDa protin was purified to near homogeneity with high yield of more than 60% using heat-treatment and anion-exchange column chromatography.The kinetic analyzes showed that Km values for this protein were caluculaed to be 0.5-1.0muM with either enzyme and suggested that the 25 kDa protein might be a candidate of natural subst5rate for these two serine/threonine kinases.5) By extensive phosphorylation, about two moles of phosphate were incorporated per mole of this protein irrespective of the protein kinase employed.The phosphorylated sites by C kinase were identified as the two serine residues locating on amino-terminal region of this protein. However, the phosphorylated sites by casein kinase II have not been conclusively identified.The data base analysis of the animo acid sequence of amino-terminal region of this 25kDa protein suggested to be a newly identified rotein.8) Amino acid comosition of this 25kDa protein indicated that this protein was similar to phosvitin, but not identical.9) The 25kDa protein was located mainly in york fraction, but this activity was detected mostly after extraction of york proteins with the buffer containing Triton X-100. This protein was also detected in other subcellular fractions and the amount was decreased according to the order of cvtosolic, microsomal and pasma membrane fractions.10) The function of this 25 kDa protein and physiological significance of the phosphorylation of this protein by C kinase and casein kinase II have been remained to be studied.