MOLECULAR-CLONING, SEQUENCE, AND EXPRESSION OF A HUMAN GDP-L-FUCOSE - BETA-D-GALACTOSIDE 2-ALPHA-L-FUCOSYL-TRANSFERASE CDNA THAT CAN FORM THE H-BLOOD GROUP ANTIGEN
MOLECULAR-CLONING, SEQUENCE, AND EXPRESSION OF A HUMAN GDP-L-FUCOSE - BETA-D-GALACTOSIDE 2-ALPHA-L-FUCOSYL-TRANSFERASE CDNA THAT CAN FORM THE H-BLOOD GROUP ANTIGEN
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DOI：
10.1073/pnas.87.17.6674
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发表时间：
1990-09-01
影响因子：
11.1
通讯作者：
LOWE, JB
中科院分区：
文献类型：
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作者：
LARSEN, RD;ERNST, LK;LOWE, JB
我们先前已使用基因转移方案分离决定GDP-L-岩藻糖：β-葡聚糖的表达的人基因组DNA片段。D-半乳糖苷2-α- L-岩藻糖基转移酶[α.（1，2）FT; EC 2.4.1.69]。尽管该片段决定了α-淀粉酶的表达。（1，2）FT，其动力学性质反映人H血型α的动力学性质。（1，2）FT，它们的确切性质仍然没有定义。我们在这里描述的分子克隆，序列和表达的人cDNA对应于这些人的基因组序列。当在COS-1细胞中表达时，该cDNA指导细胞表面H结构和同源α的表达。（1，2）具有与人H血型α类似的性质的FT活性（1，2）FT.该cDNA序列预测了具有II型跨膜糖蛋白特征的365个氨基酸的多肽，其结构域结构类似于其它糖基转移酶的结构域结构，但与这些或其它已知蛋白质没有显著的一级序列相似性。为了直接证明该cDNA编码α.（1，2）FT，预测为高尔基体驻留的COOH末端结构域，在COS-1细胞中表达为催化活性的分泌的可溶性蛋白A融合肽。Southern印迹分析表明，该cDNA鉴定了与人类19号染色体上的H位点同线的DNA序列。这些结果有力地表明，这种克隆的α。（1，2）FT cDNA代表人H血型基因座的产物。
We have previously used a gene-transfer scheme to isolate a human genomic DNA fragment that determines expression of a GDP-L-fucose:.beta.-D-galactoside 2-.alpha.-L-fucosyltransferase [.alpha.(1,2)FT; EC 2.4.1.69]. Although this fragment determined expression of an .alpha.(1,2)FT whose kinetic properties mirror those of the human H blood group .alpha.(1,2)FT, their precise nature remained undefined. We describe here the molecular cloning, sequence, and expression of a human cDNA corresponding to these human genomic sequences. When expressed in COS-1 cells, this cDNA directs expression of cell surface H structures and a cognate .alpha.(1,2)FT activity with properties analogous to the human H blood group .alpha.(1,2)FT. The cDNA sequence predicts a 365-amino acid polypeptide characteristic of a type II transmembrane glycoprotein with a domain structure analogous to that of other glycosyltransferases but without significant primary sequence similarity to these or other known proteins. To directly demonstrate that the cDNA encodes an .alpha.(1,2)FT, the COOH-terminal domain predicted to be Golgi-resident was expressed in COS-1 cells as a catalytically active, secreted, and soluble protein A fusion peptide. Southern blot analysis showed that this cDNA identifies DNA sequences syntenic to the human H locus on chromosome 19. These results strongly suggest that this cloned .alpha.(1,2)FT cDNA represents the product of the human H blood group locus.