Acceptor substrate discrimination in phosphatidyl-myo-inositol mannoside synthesis: structural and mutational analysis of mannosyltransferase Corynebacterium glutamicum PimB'.
Acceptor substrate discrimination in phosphatidyl-myo-inositol mannoside synthesis: structural and mutational analysis of mannosyltransferase Corynebacterium glutamicum PimB'.
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DOI：
10.1074/jbc.m110.165407
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发表时间：
2010-11-26
期刊：
影响因子：
--
通讯作者：
Fütterer K
中科院分区：
文献类型：
--
作者：
Batt SM;Jabeen T;Mishra AK;Veerapen N;Krumbach K;Eggeling L;Besra GS;Fütterer K
结核分枝杆菌在人类体内的长期存活与其复杂的细胞壁糖脂的免疫调节潜力有关，其中包括磷脂酰肌醇甘露糖苷(PIM)系列以及相关的脂甘露聚糖和脂阿拉伯甘露糖糖结合物。PIM的生物合成是由一组胞内α-甘露糖基转移酶启动的，催化糖基从激活的糖供体Gdp-α-d-甘露糖基转移到受体磷脂酰肌醇(PI)。在这里，我们报道了甘露糖基转移酶谷氨酸棒杆菌PimB‘与核苷酸的络合物的晶体结构，分辨率为2.0Å。PimB‘选择性地将甘露醇结合到PI的肌醇部分的6-OH上。两种晶型和Gdp-α-d-甘露糖结合络合物揭示了活性中心的核苷酸构象和结构框架的灵活性。结构比较，糖受体的对接，以及定点突变对N-末端结构域中保守的Asp残基的区域选择性，迫使正确的肌醇羟基呈现给糖供体。
Long term survival of the pathogen Mycobacterium tuberculosis in humans is linked to the immunomodulatory potential of its complex cell wall glycolipids, which include the phosphatidylinositol mannoside (PIM) series as well as the related lipomannan and lipoarabinomannan glycoconjugates. PIM biosynthesis is initiated by a set of cytosolic α-mannosyltransferases, catalyzing glycosyl transfer from the activated saccharide donor GDP-α-d-mannopyranose to the acceptor phosphatidyl-myo-inositol (PI) in an ordered and regio-specific fashion. Herein, we report the crystal structure of mannosyltransferase Corynebacterium glutamicum PimB′ in complex with nucleotide to a resolution of 2.0 Å. PimB′ attaches mannosyl selectively to the 6-OH of the inositol moiety of PI. Two crystal forms and GDP- versus GDP-α-d-mannopyranose-bound complexes reveal flexibility of the nucleotide conformation as well as of the structural framework of the active site. Structural comparison, docking of the saccharide acceptor, and site-directed mutagenesis pin regio-selectivity to a conserved Asp residue in the N-terminal domain that forces presentation of the correct inositol hydroxyl to the saccharide donor.