ENZYMATIC PEPTIDE-SYNTHESIS IN LOW WATER-CONTENT SYSTEMS - PREPARATIVE ENZYMATIC-SYNTHESIS OF [LEU]-ENKEPHALIN AND [MET]-ENKEPHALIN DERIVATIVES

被引:38
作者
CLAPES, P
TORRES, JL
ADLERCREUTZ, P
机构
[1] CSIC, CID, PROT CHEM & BIOCHEM UNIT, E-08034 BARCELONA, Spain
[2] LUND UNIV, CTR CHEM, DEPT BIOTECHNOL, S-22100 LUND, Sweden
关键词
D O I
10.1016/0968-0896(95)00019-D
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
A novel total enzymatic synthesis of [Leu]- and [Met]-enkephalin derivatives was accomplished in low-water content systems at a preparative scale. alpha-Chymotrypsin, papain, thermolysin and bromelain adsorbed on Celite were used as catalysts. Organic solvents such as acetonitrile and ethyl acetate with small amounts of buffer added or at specific water activity were used as reaction media. Simple readily available amino acid ester derivatives were used as starting building blocks. This feature allowed the possibility of using the products in one step directly as acyl-donor ester, without any chemical or enzymatic modification, in the next enzymatic coupling. The optimal strategy for the synthesis of the enkephalin derivatives was different depending on the carboxy terminal group. The preparation of the carboxy-terminal amide derivatives (R-Tyr-Gly-Gly-Phe-Leu[Met]-NH2) was achieved via 4 + 1 fragment condensation catalyzed by alpha-chymotrypsin. The carboxy-terminal ethyl ester derivatives (R-Tyr-Gly-Gly-Phe-Leu[Met]-OEt) were obtained via 2 + 3 condensation catalyzed by bromelain, a quite unusual protease for peptide synthesis but more effective than papain in this coupling. Both syntheses were carried out in four enzymatic steps and one or two chemical deprotection steps routinely used in peptide synthesis. The overall yields of pentapeptide derivatives were between 40-54% of pure product.
引用
收藏
页码:245 / 255
页数:11
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