Isolation and characterization of the Saccharomyces cerevisiae DPP1 gene encoding diacylglycerol pyrophosphate phosphatase

被引:111
作者
Toke, DA
Bennett, WL
Dillon, DA
Wu, WI
Chen, XM
Ostrander, DB
Oshiro, J
Cremesti, A
Voelker, DR
Fischl, AS
Carman, GM [1 ]
机构
[1] Rutgers State Univ, Cook Coll, New Jersey Agr Expt Stn, Dept Food Sci, New Brunswick, NJ 08903 USA
[2] Natl Jewish Ctr Immunol & Resp Med, Lord & Taylor Lab Lung Biochem, Denver, CO 80206 USA
[3] Natl Jewish Ctr Immunol & Resp Med, Anna Perahia Adatto Clin Res Ctr, Denver, CO 80206 USA
[4] Univ Rhode Isl, Dept Food Sci & Nutr, W Kingston, RI 02892 USA
关键词
D O I
10.1074/jbc.273.6.3278
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
Diacylglycerol pyrophosphate (DGPP) is involved in a putative novel lipid signaling pathway, DGPP phosphatase (DGPP phosphohydrolase) is a membrane-associated 34-kDa enzyme from Saccharomyces cerevisiae which catalyzes the dephosphorylation of DGPP to yield phosphatidate (PA) and then catalyzes the dephosphorylation of PA to yield diacylglycerol, Amino acid sequence information derived from DGPP phosphatase was used to identify and isolate the DPP1 (diacylglycerol pyrophosphate phosphatase) gene encoding the enzyme. Multicopy plasmids containing the DPP1 gene directed a 10-fold overexpression of DGPP phosphatase activity in S. cerevisiae. The heterologous expression of the S. cerevisiae DPP1 gene in Sf-9 insect cells resulted in a 500-fold overexpression of DGPP phosphatase activity over that expressed in wild-type S. cerevisiae. DGPP phosphatase possesses a Mg2+-independent PA phosphatase activity, and its expression correlated with the overexpression of DGPP phosphatase activity in S. cerevisiae and in insect cells. DGPP phosphatase was predicted to be an integral membrane protein with six transmembrane-spanning domains. The enzyme contains a novel phosphatase sequence motif found in a superfamily of phosphatases. A dpp1 Delta mutant was constructed by deletion of the chromosomal copy of the DPP1 gene. The dpp1 Delta mutant was viable and did not exhibit any obvious growth defects. The mutant was devoid of DGPP phosphatase activity and accumulated (4-fold) DGPP. Analysis of the mutant showed that the DPP1 gene was not responsible for all of the Mg2+-independent PA phosphatase activity in S. cerevisiae.
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收藏
页码:3278 / 3284
页数:7
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