Highly efficient chemoenzymatic synthesis of α-galactosyl epitopes with a recombinant α(1→3)-galactosyltransferase
Highly efficient chemoenzymatic synthesis of α-galactosyl epitopes with a recombinant α(1→3)-galactosyltransferase
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DOI：
10.1021/ja9808898
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发表时间：
1998-07-15
影响因子：
15
通讯作者：
Wang, PG
中科院分区：
文献类型：
--
作者：
Fang, JW;Li, J;Wang, PG
α -半乳糖表位是具有α 1-3Gal末端的碳水化合物结构。动物细胞表面的这些抗原表位与人血清中抗α -半乳糖抗体的相互作用被认为是异种移植中抗体介导的超急性排斥反应的主要原因。本报告描述了一种基于重组α(1—bbbb3)-半乳糖转移酶（α 1,3- galt）的高效化学酶方法，用于合成异种活性α -半乳糖表位，这在异种移植和免疫治疗研究中是非常需要的。将截断的牛α 1,3- galt（80-368）克隆到pET15b载体中，随后转化为大肠杆菌BL21菌株。该表达体系高效、大规模地产生了具有高特异性活性的可溶性重组酶。利用该重组酶合成了多种α（1—>3）半乳糖基化表位。以一种独特的方式，α -半乳糖五糖通过一锅，两步酶合成与原位辅助因子再生合成。
alpha-Galactosyl epitopes are carbohydrate structures bearing a Gal alpha 1-3Gal beta terminus. The interaction of these epitopes on the surface of animal cells with anti-alpha-galactosyl antibodies in human serum is believed to be the main cause in antibody-mediated hyperacute rejection in xenotransplantation. This report describes an efficient chemoenzymatic approach based on the use of recombinant alpha(1 --> 3)-galactosyltransferase (alpha 1,3-GalT) for the synthesis of xenoactive alpha-galactosyl epitopes, which are highly desired in the research of xenotransplantation and immunotherapy. A truncated bovine alpha 1,3-GalT (80-368) was cloned into the pET15b vector-and Subsequently transformed into E. coli BL21 strain. This expression system efficiently produced the soluble recombinant enzyme on a large scale with highly specific activity. A variety of alpha(1 --> 3)galactosylated epitopes were synthesized using such a recombinant enzyme. In a unique fashion, alpha-galactosyl pentasaccharide was synthesized via a one-pot, two-step enzymatic synthesis with in situ cofactor regeneration.