Mono(ADP-ribosyl)ation reaction and its functions in primary cultured hepatocytes
| Project/Area Number |
60570112
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
General medical chemistry
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| Research Institution | Depertment of biochemistry, Shimane medical university |
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Principal Investigator |
TANIGAWA Yoshinori Department of biochemistry, Shimane medical university assistant professor, 医学部, 助教授 (60084860)
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| Co-Investigator(Kenkyū-buntansha) |
MISHIMA Kouichi Department of biochemistry, Shimane medical university assistant, 医学部, 助手 (90181875)
TUCHIYA Mikako Department of biochemistry, Shimane medical university assistant, 医学部, 助手 (90188582)
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| Project Period (FY) |
1985 – 1986
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| Project Status |
Completed (Fiscal Year 1986)
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| Budget Amount *help |
¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 1986: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1985: ¥800,000 (Direct Cost: ¥800,000)
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| Keywords | Mono(ADP-ribose) / Protein phosphorylation / Membran proteins / 肝初代培養 |
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| Research Abstract |
(1)Phosphorylation of Kemptide (Leu-Arg-Arg-Ala-Ser-Leu-Gly) by cyclic AMP- or <Ca^(2+)> , phospholipid-dependent protein kinase occured the serine residue,and also phosphorylation of Reverse-kemptide (Gly-Leu-Ser-Ala-Arg-Arg-Leu) by <Ca^(2+)> ,phospholipid-dependent protein kinase occured at the serine residue, but Kemptide dose not served as substrate for cyclic AMP-dependent protein kinase.
(2)ADP-ribosylation of Kemptide or Reverse-kemptide by hen liver nuclear ADP-ribosyltransferase occured at the two arginine residues sequenced at the <NH_2> -terminal(Kemptide)or COOH-terminal(Reverse-kemptide) side of the phosphate-accepting serine residue.
(3)Phosphorylations of Kemptide and Reverse-kemptide by cyclic AMP-dependent and <Ca^(2+)> , phospholipid-de-pendent protein kinase, respectively,were markedly reduced when both synthetic heptapeptides were ADP-ribosylated. Kinetic studies of phosphorylation revealed that ADP-ribosylated both synthetic peptides were a linear competitive inhibitor of each synthetic peptide and a linear noncompetitive inhibitor of ATP.
(4)Primary cultured hepatocite were labeled with [ <^(32)P> ]orthophosphate and [ <^3H> ]adenosine, respectively.Polyacrylamide gel analyses of membrane preparation from these cells revealed that over ten proteins are ADP-ribosylated and at least eight proteins are phosphorylated. Four proteins has molecular weight of 85,000, 60,000, 42,000, and 20,000 proved to be effectve acceptor for both modifications.
(5)When 20 mM nicotinamide was added into the praimary culture medium, 30 % decrease of total protein phosphorylation were observed. The phosphorylation reduced was predominantly observed to two proteins which has molecular weight of 60,000 and 42,000.
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Research Products
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