Structural basis of ordered binding of donor and acceptor substrates to the retaining glycosyltransferase, α-1,3-galactosyltransferase
Structural basis of ordered binding of donor and acceptor substrates to the retaining glycosyltransferase, α-1,3-galactosyltransferase
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DOI：
10.1074/jbc.m202631200
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发表时间：
2002-08-02
影响因子：
4.8
通讯作者：
Acharya, KR
中科院分区：
文献类型：
--
作者：
Boix, E;Zhang, YN;Acharya, KR
牛 α-1,3-半乳糖基转移酶 (α3GT) 催化 α-半乳糖 (α-Gal) 表位的合成，这是天然人类抗体的靶标。它代表一个酶家族，包括组织血型 A 和 B 转移酶，可催化未知机制的保留糖基转移反应。对具有有限分辨率和相当大无序的晶体形式的 α3GT 的初步研究表明可能形成 β-半乳糖基-酶共价中间体（Gstinel, L. N.、Bignon, C.、Misra, A. K.、Hindsgaul, O.、Shaper, J. H. 和 Joziasse, D. H. (2001) EMBO J. 20, 638-649)。描述了 alpha3GT 与供体底物 UDP-半乳糖、UDP 葡萄糖和两种受体底物乳糖和 N-乙酰乳糖胺的复合物的高度有序结构，分辨率高达 1.46 埃。结构和量热结合研究表明，供体和受体底物必须有序结合，与供体底物诱导的构象变化有关，并且 UDP 直接参与受体结合。单糖-UDP 键在含有 UDP-半乳糖和 UDP 葡萄糖的结构中裂解，产生含有埋藏的 β-半乳糖和 α-葡萄糖的非共价复合物。这些单糖的位置和分子模型表明扭曲构象的 UDP-半乳糖的结合可能在 alpha3GT 的催化机制中很重要。
Bovine alpha-1,3-galactosyltransferase (alpha3GT) catalyzes the synthesis of the a-galactose (alpha-Gal) epitope, the target of natural human antibodies. It represents a family of enzymes, including the histo blood group A and B transferases, that catalyze retaining glycosyltransfer reactions of unknown mechanism. An initial study of alpha3GT in a crystal form with limited resolution and considerable disorder suggested the possible formation of a beta-galactosyl-enzyme covalent intermediate (Gastinel, L. N., Bignon, C., Misra, A. K., Hindsgaul, O., Shaper, J. H., and Joziasse, D. H. (2001) EMBO J. 20, 638-649). Highly ordered structures are described for complexes of alpha3GT with donor substrate, UDP-galactose, UDPglucose, and two acceptor substrates, lactose and N-acetyllactosamine, at resolutions up to 1.46 Angstrom. Structural and calorimetric binding studies suggest an obligatory ordered binding of donor and acceptor substrates, linked to a donor substrate-induced conformational change, and the direct participation of UDP in acceptor binding. The monosaccharide-UDP bond is cleaved in the structures containing UDP-galactose and UDPglucose, producing non-covalent complexes containing buried beta-galactose and alpha-glucose. The location of these monosaccharides and molecular modeling suggest that binding of a distorted conformation of UDP-galactose may be important in the catalytic mechanism of alpha3GT.