Cloning and expression of human core 1 β1,3-galactosyltransferase
Cloning and expression of human core 1 β1,3-galactosyltransferase
复制标题
DOI：
10.1074/jbc.m109060200
复制
发表时间：
2002-01-04
影响因子：
4.8
通讯作者：
Canfield, WM
中科院分区：
文献类型：
--
作者：
Ju, TZ;Brewer, K;Canfield, WM
共同的核心10-聚糖结构Galbeta1-> 3GalNAc-R是动物细胞表面和分泌的糖蛋白中许多延伸的粘蛋白型0-聚糖结构的前体。核心1是通过核心1 β 3-半乳糖基转移酶（核心1 β 3-Gal-T）将Gal从UDP-Gal转移至GalNAcalpha1-R而合成的。来自纯化的大鼠核心1 β 3-Gal-T的氨基酸序列（Ju，T.，卡明斯河D、和坎菲尔德，W. M.（2002）J.Biol.Chem.277，169 - 177）用于鉴定人表达序列标签数据库中的核心1 β 3-Gal-T序列。通过对表达的序列标签进行测序并进行cDNA末端的5 ′-快速扩增来确定1794-bp的人核心1 β 3-Gal-T cDNA序列。核心1 β 3-Gal-T预测了363个氨基酸的II型跨膜蛋白。在人293 T细胞中表达全长和表位标记的可溶性形式的推定酶产生核心1 β 3-Gal-T活性，其将半乳糖从UDP-Gal转移到GalNAcalpha1-O-苯基，并且具有Thr连接的GalNAc的合成糖肽和产物显示具有核心I结构。北方分析表明核心1 β 3-Gal-T在组织中广泛表达，其中在肾脏、心脏、胎盘和肝脏中占主导地位。从大鼠、小鼠、果蝇和秀丽隐杆线虫中鉴定并克隆了高度同源的cDNA，表明该酶广泛分布于后生动物中。核心1 β 3-Gal-T序列与先前描述的β 3-半乳糖基转移酶中发现的保守序列具有最小的同源性，表明该酶与已知的β 3-半乳糖基转移酶家族仅有远亲关系。
The common core 1 O-glycan structure Galbeta1-->3GalNAc-R is the precursor for many extended mucin-type O-glycan structures in animal cell surface and secreted glycoproteins. Core 1 is synthesized by the transfer of Gal from UDP-Gal to GalNAcalpha1-R by core 1 beta3-galactosyltransferase (core 1 beta3-Gal-T). Amino acid sequences from purified rat core 1 beta3-Gal-T (Ju, T., Cummings, R. D., and Canfield, W. M. (2002) J. Biol. Chem. 277, 169-177) were used to identify the core 1 beta3-Gal-T sequences in the human expressed sequence tag data bases. A 1794-bp human core 1 beta3-Gal-T cDNA sequence was determined by sequencing the expressed sequence tag and performing 5'-rapid amplification of cDNA ends. The core 1 beta3-Gal-T predicts a 363-amino acid type II transmembrane protein. Expression of both the full-length and epitope-tagged soluble forms of the putative enzyme in human 293T cells generated core 1 beta3-Gal-T activity that transferred galactose from UDP-Gal to GalNAcalpha1-O-phenyl, and a synthetic glycopeptide with Thr-linked GalNAc and the product was shown to have the core I structure. Northern analysis demonstrated widespread expression of core 1 beta3-Gal-T in tissues with a predominance in kidney, heart, placenta, and liver. Highly homologous cDNAs were identified and cloned from rat, mouse, Drosophila melanogaster, and Caenorhabditis elegans, suggesting that the enzyme is widely distributed in metazoans. The core 1 beta3-Gal-T sequence has minimal homology with conserved sequences found in previously described beta3-galactosyltransferases, suggesting this enzyme is only distantly related to the known beta3-galactosyltransferase family.