YEAST DOLICHYL-PHOSPHOMANNOSE SYNTHASE - RECONSTITUTION OF ENZYME-ACTIVITY WITH PHOSPHOLIPIDS

被引:21
作者
SCHUTZBACH, JS [1 ]
ZIMMERMAN, JW [1 ]
机构
[1] MIT,CTR CANC RES,CAMBRIDGE,MA 02139
来源
BIOCHEMISTRY AND CELL BIOLOGY-BIOCHIMIE ET BIOLOGIE CELLULAIRE | 1992年 / 70卷 / 06期
关键词
DOLICHYL PHOSPHATE; MANNOSE; SYNTHASE; TRANSPORT; LIPOSOMES;
D O I
10.1139/o92-070
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 activity of purified recombinant yeast dolichyl-phosphomannose synthase (EC 2.4.1.83) was assessed following reconstitution of the enzyme with phospholipids. The yeast synthase, similar to the mammalian enzyme, was active when reconstituted with phosphatidylethanolamine dispersions but had little (less than 5%) activity in stable phosphatidylcholine bilayers. The enzyme was activated by adding increasing amounts of diacylglycerol to phospholipid bilayers, suggesting that activity of the yeast enzyme was dependent on lipid phase properties rather than specific phospholipids. The synthase could also be reconstituted as an active enzyme in bilayers prepared with a commercial crude lipid preparation containing 40% phosphatidylcholine, but at a rate 10% of that occurring in phosphatidylethanolamine. Vesicles composed of the 40% phosphatidylcholine lipid mixture, dolichyl phosphate, and enzyme were leaky in the presence of divalent cations, and dolichyl-phosphomannose synthase did not appear to catalyze the translocation of dolichyl phosphomannose across membranes at a catalytically significant rate under the assay conditions employed.
引用
收藏
页码:460 / 465
页数:6
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