| Summary: | 碩士 === 國立臺灣大學 === 生化科學研究所 === 83 === Agkistrodon acutus venom contains a variety of proteins that
may effect vertebrate blood coagulation and platelet funcion.
Two novel sanke venom proteins with antiplatelet activity, agki-cetin-A and agkicetin-B, were purified from Agkistrodon acutus (i.e., Deinagkistrodon acutus) by cationic exchanger chromato-graphy and RP-HPLC. Both agkicetin isoforms are heterodimeric proteins with disulfide bond linkages. Determination of the apparent molecular weight of agkicetin was performed on SDS-gel electrophoresis. The apparent molecular weight of agkicetin-A is 26KDa, and that of agkicetin-B is 25KDa; the reduced two sub- units are 15KDa and 14KDa, respectively. Amino acid analysis showed that agkicetin isoforms are not only rich in Asp/Asp, Glu /Gln, Ser and Gly, but also highly homologous. The N-terminal amino acid sequence of agkicetin isoforms showed 64~70% identity to each other.
Agkicetin isforms did not bind coagulation factor IX and
thrombin apparently. Both of agkicetin isoforms behaved as a
potent antagonist of von Willebrand factor (vWF) - induced
platelet agglutination and bound specifically to glycoprotein Ib
(GPIb) of fixed human platelets. However, the antiplatelet acti-
vity of agkicetin isoforms was very different. Agkicetin-A was
85 times more active than agkicetin-B in inhibiting bovine vWF-
incuced platelet agglutination. We also showed that oxidation of
methionine residues in agkicetin-A with chloramine T decreased
its inhibitory activity toward bovine vWF induced platelet
agglutination and its binding to GPIb.
The secondary structure of agkicetin isoforms was studied by circular dichrosim, and the result showed that agkicetin
isoforms are highly homologous. This preliminary study suggests
that agkicetin isoforms may be utilized as a research tool to
develop new antithrombotic agents. In addition, both isoforms
would help to elucidate the structure-function relationship of
this family of antiplatelet proteins.
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