Zhu X Y, Lehrman M A
Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas 75235.
J Biol Chem. 1990 Aug 25;265(24):14250-5.
UDP-GlcNAc:dolichol phosphate N-acetylglucosamine-1-phosphate transferase (GPT) catalyzes the initial reaction required for synthesis of dolichol-P-P-oligosaccharides. We report here on the sequence and expression of a full-length cDNA clone encoding hamster GPT. The cDNA predicts a protein of 408 amino acid residues including 10 hydrophobic segments. Several portions of the hamster GPT sequence constituting one-third of the protein have 60% or greater identity with yeast GPT, and one-half of the conserved sequence falls within the hydrophobic segments. In addition, hamster GPT has two copies of a putative dolichol recognition sequence recently identified in three yeast enzymes that interact with dolichol. The protein lacks KDEL or DEKKMP-type carboxyl-terminal ER sorting sequences. When expressed in COS-1 cells, the cDNA causes a 5-7-fold increase of GPT activity in membrane fractions. The activity was completely inhibitable by tunicamycin, and the primary product was shown to be GlcNAc-pyrophosphoryldolichol. This cDNA represents the first enzyme of the dolichol-oligosaccharide biosynthetic pathway to be cloned from a vertebrate source and demonstrates structural homology between the enzymes of the yeast and mammalian pathways.
UDP-葡萄糖胺：磷酸多萜醇N-乙酰葡糖胺-1-磷酸转移酶（GPT）催化合成多萜醇-焦磷酸-寡糖所需的起始反应。我们在此报告编码仓鼠GPT的全长cDNA克隆的序列和表达情况。该cDNA预测编码一个含有408个氨基酸残基的蛋白质，其中包括10个疏水片段。仓鼠GPT序列中构成该蛋白质三分之一的几个部分与酵母GPT有60%或更高的同源性，且保守序列的一半位于疏水片段内。此外，仓鼠GPT有两个最近在三种与多萜醇相互作用的酵母酶中鉴定出的假定多萜醇识别序列拷贝。该蛋白质缺乏KDEL或DEKKMP型羧基末端内质网分选序列。当在COS-1细胞中表达时，该cDNA使膜组分中的GPT活性增加5至7倍。衣霉素可完全抑制该活性，且主要产物被证明是GlcNAc-焦磷酸化多萜醇。该cDNA代表了从脊椎动物来源克隆的多萜醇-寡糖生物合成途径的首个酶，并证明了酵母和哺乳动物途径的酶之间存在结构同源性。
