소뇌에서 Protein carboxyl methyltransferase의 정제 및 특성에 관한 연구

Abstract

Protein carboxyl methyltransferase(S-adenosyl-methionine;protein-carboxylo-methyltransferase,E. C. 2.1.1.77) of which methvlesterified free carboxyl group of protein substrate using S-adenosyl-L-methionine as the methyl donor has been prified from bovine brain approximatly 1,061-fold with a yield of 25% The enzymes were purified by following steps:70% ammonium sulfate,DEAE-cellulose chromatography, SAH-sepharose 4 B affinity chromatography. PCM from bovin braine was found to exist as three isoenzymes that could be seperated by tow-dimensional electrophoresis and DEAE - 5PW anion exchang high-performance liquid chromtograph. PⅠ Values of bovine brain PCM were 5.8, 6.2, and 6.25 by two-dimensional electrophoresis. The moleclar weights of isoenzymes of which Pl values were'5.8,6,2 and 6,25 by two-dimensional electrophoresis were-28,500,28,000,28,000 daltons respectivily. Mixture of the PCH isoenzyme purified by SAH-affinity chromatograph were subjected to high-perfo-rmance liquid chromtography on a DEAE - 5PW anion exchange colum. Result of HPLC analysis indicated that bovine brain PCM has distinct three isoenzymes.;Methyl donor로 S - adenosyl - L - methionine을 사용해서 기질단백질의 free carboxyl group을 methylester화 시키는 반응을 촉매하는 Protein Carboxyl Methyltransferase (PCM)를 소뇌에서 25 %의 수득률료 1,061 배 정제하였다. 소뇌의 cytosol 를 70 % ammonium sulfate, DEAE - cellulose chromatography, SAH - sepharose 4B affinity chromatograhy방법으로 정제하였으며 정제된 효소용액을 electrophoresis한 결과 PCM의 PI가 5.80, 6.20, 6.25로 밝혀졌으며, PI가 5.80인 isoenzyme의 분자량은 28,500 dalton이었고, PI가 6.20, 6.25인 isoenzyme의분자량은 모두 28,000 dalton이었다. 또한 정제된효소용 액중에 존재하는 세 종류의 PCM isoenzyme을 음이온 교환 HPLC로 분리하였다.