The amino acids involved in the distinct carbohydrate specificities between macrophage galactose-type C-type lectins 1 and 2 (CD301a and b) of mice
The amino acids involved in the distinct carbohydrate specificities between macrophage galactose-type C-type lectins 1 and 2 (CD301a and b) of mice
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DOI：
10.1016/j.bbagen.2007.10.017
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发表时间：
2008-02-01
影响因子：
3
通讯作者：
Irimura, Tatsuro
中科院分区：
文献类型：
--
作者：
Oo-puthinan, Sarawut;Maenuma, Keisuke;Irimura, Tatsuro
采用正面亲和层析法比较了小鼠巨噬细胞半乳糖型C型凝集素1（MGL 1/CD 301 a）和2（MGL 2/CD 301 b）与各种寡糖的结合特异性。在111种测试的寡糖中，MGL 1优先结合含有刘易斯（X）（Le（X））三糖苷的寡糖，而MGL 2优先结合球糖苷Gb 4。通过在大肠杆菌中制备并用半乳糖-琼脂糖凝胶纯化的可溶性重组碳水化合物识别结构域（CRD）中的位置61、89、97、100、110-113、115、124和125处进行成对定点突变，研究了优先结合的重要氨基酸。MGL 1 CRD上61、89、111和125位的瓦尔、Ala、Thr和Phe突变导致Le（X）结合减少。MGL 2 CRD在第61、89、115和125位的Leu、Arg、Arg和Tyr处的突变涉及对β-GalNAc的偏好。在MGL 2的Arg 89 Ala和Arg 89 Ala/Ser 111 Thr突变体中观察到Le（X）结合。MGL 1的Ala 89 Arg和Ala 89 Arg/Pro 115 Arg突变体显示β-GalNAc结合。分子模型说明了MGL 2 CRD中Leu 61、Arg 89和His 109与GalNAc的潜在直接分子相互作用。(C)2007 Elsevier B. V.保留所有权利。
Binding specificities of mouse macrophage galactose-type C-type lectin 1 (MGL1/CD301a) and 2 (MGL2/CD301b) toward various oligosaccharides were compared by frontal affinity chromatography. MGL1 preferentially bound oligosaccharides containing Lewis(X) (Le(X)) trisaccharides among 111 oligosaccharides tested, whereas MGL2 preferentially bound globoside Gb4. The important amino acids for the preferential bindings were investigated by pair-wise site-directed mutagenesis at positions 61, 89, 97, 100, 110-113, 115, 124, and 125 in the soluble recombinant carbohydrate recognition domains (CRD) prepared in Escherichia coli and purified with galactose-Sepharose. Mutations of Val, Ala, Thr, and Phe at positions 61, 89, 111 and 125 on MGL1 CRD caused reductions in Le(X) binding. Mutations of MGL2 CRD at Leu, Arg, Arg, and Tyr at positions 61, 89, 115 and 125 were implicated in the preference for beta-GalNAc. Le(X) binding was observed with MGL2 mutants of Arg89Ala and Arg89Ala/Ser111 Thr. MGL1 mutants of Ala89Arg and Ala89Arg/Pro115Arg showed beta-GalNAc bindings. Molecular modeling illustrated potential direct molecular interactions of Leu61, Arg89, and His109 in MGL2 CRD with GalNAc. (C) 2007 Elsevier B.V. All rights reserved.