PSEUDOMONAS-GLUMAE LIPASE - INCREASED PROTEOLYTIC STABILITY BY PROTEIN ENGINEERING

被引:30
作者
FRENKEN, LGJ
EGMOND, MR
BATENBURG, AM
VERRIPS, CT
机构
[1] Unilever Research Laboratorium, Vlaardingen, AT, 3133
来源
PROTEIN ENGINEERING | 1993年 / 6卷 / 06期
关键词
LIPASE; PROTEIN ENGINEERING; PSEUDOMONAS-GLUMAE; PROTEOLYTIC CLEAVAGE SITE; SUBTILISIN;
D O I
10.1093/protein/6.6.637
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
The feasibility of stabilizing proteins towards proteolytic degradation was explored by engineering the primary proteolytic cleavage site(s). This novel approach does not require information on the 3-D structure of the native enzyme. As a model system, the extracellular lipase of Pseudomonas glumae was chosen, which is sensitive towards degradation by subtilisin-type proteases. The primary proteolytic cleavage in the lipase appeared to be located between amino acids serine 153 and histidine 154. Since subtilisins are known to show a preference towards amino acid residues surrounding the scissile bond, non-preferred amino acids were introduced in this area. Two concepts were tested: the introduction of arginine or glutamate residues (charge concept) and the introduction of proline residues (proline concept). Although the mutant lipases produced according to either of these concepts were still cleaved in the same area, they showed a considerably increased stability towards proteolytic degradation.
引用
收藏
页码:637 / 642
页数:6
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