| Budget Amount *help |
¥6,800,000 (Direct Cost: ¥6,800,000)
Fiscal Year 1995: ¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 1994: ¥4,400,000 (Direct Cost: ¥4,400,000)
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| Research Abstract |
In a porcine aorta extract, we observed two protein kinase activities which specifically phosphorylate the 204-kDa heavy chain isoform of aorta myosin in the absence of conventional kinase activators. We referred to these two protein kinases, eluted at 0.15 and 0.2 M KCl from a DEAE-column, as myosin kinases I (MKI) and II (MKII) , respectively. The phosphorylation site for MKI was determined using a purified phosphopeptide derived from porcine aorta myosin phosphorylated with MKI.By comparison with the deduced amino acid sequence for smooth muscle myosins, the site corresponded to a Ser located at 3 amino acids upstream from a Pro, the putative end of the alpha-helical segment of the 204-kDa heavy chain tail. A homologous Ser is only present in smooth muscle myosins, i. e. not in nonmuscle myosins. MKI was purified 130-fold, but not separated from a kinase activity phosphorylating Ser1 or Ser2 in the 20-kDa regulatory light chain of aorta myosin. In contrast, MKII was purified to near homogeneity. MKII phosphorylated the porcine aorta myosin heavy chain at a Ser 19 amino acids downstream from the MKI site. The amino acid sequence around the Ser shared a consensus sequence of the phosphorylation site for casein kinase II and was homologous to that reported for bovine aorta myosin (Kelley, C.A., and Adelstein, R.S.(1990) J.Biol.Chem.265,17876-17882) . MKII was identified as a multifunctional protein kinase, casein kinase II.
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