| Co-Investigator(Kenkyū-buntansha) |
KAWAGUCHI Kiichiro Lecture, Dept.of Pharmacology, School of Pharmaceutical Sciences, Kitasato Unive, 薬学部, 講師 (10146334)
TANAKA Yuetsu Associate Professor, Det.of Immunology, School of Sciences, Kitasato University, 理学部, 助教授 (30163588)
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| Research Abstract |
The project study was done to investigate (i) purification of the specific binding proteins (gbPs) of anti-inflammatory egent, glycyrrhizin, from GL-sensitive mammalian cells, (ii) identification of gbPs by determination of their N-terminal partial amino acid sequences, and (iii) kinetics of the GL-induced selective inhibition of their physiological activities in vitro.
To purify the gbPs from GL-sensitive mammalian cells or mouse tissues (liver and spleen) , a GL-affinity column was prepared. By using this GL-affinity column, several gbPs were purified to homogenous polypeptides after partial purification of the 1.5 M NaC1 clude extracts by combination with CM-cellulose column cromatography and gel filtration on a Superdex 200pg column (HPLC) . Determination of their N-termina partial amino acid sequences revealed that (i) arachidonate cascade enzymes, such as phospholipase A_2 (PLA_2) , 5'-1ipoxygenase (5'-LOX) and lipocortin I,are identified as gbPs ; (ii) serine protease (p32) , hyaluronidase (p55) comperiment C3 and immunoglobulin (Igg) are identified as gbPs in human serum ; and (iii) 14 kDa secetory PLA_2 (14 kDa sPLA_2 ) and serine protease are identified as gbPs in the synovial fluids of patients with rheumatonoid arthritis. Furthermore, kenitic studies revealed that GL,but not GA,at low levels (1-5muM) inhibits the-physiological activities of these gbPs and also selectively inhibits the CK-II-mediated activation of these gbPs in vitro.
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