Recognition of Heteropolysaccharide Alginate by Periplasmic Solute-Binding Proteins of a Bacterial ABC Transporter
Recognition of Heteropolysaccharide Alginate by Periplasmic Solute-Binding Proteins of a Bacterial ABC Transporter
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DOI：
10.1021/bi300194f
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发表时间：
2012-05-01
期刊：
影响因子：
2.9
通讯作者：
Murata, Kousaku
中科院分区：
文献类型：
--
作者：
Nishitani, Yu;Maruyama, Yukie;Murata, Kousaku
藻酸盐是由β-D-甘露糖醛酸（M）和α-L-古洛糖醛酸（G）组成的杂多糖。革兰氏阴性菌鞘氨醇单胞菌A1通过周质将藻酸盐直接掺入细胞质中。可溶性结合蛋白（AlgQ 1和AlgQ 2）依赖性ABC转运蛋白（AlgM 1-AlgM 2/AlgS-AlgS）。两个结合蛋白，至少有四个亚位点强烈识别的非还原性末端残基的藻酸盐在亚位点1。在这里，我们展示了菌株A1溶质结合蛋白对藻酸盐中存在的M和G的广泛底物偏好，并通过与饱和和不饱和藻酸盐低聚糖复合的四种AlgQ 1结构的X射线晶体学证明了杂多糖识别结合蛋白中的结构决定因素。不同M/G比的藻酸盐被菌株A1细胞同化并与AlgQ 1和AlgQ 2结合。寡糖结合形式的晶体结构显示，除了AlgQ 1和不饱和寡糖之间的相互作用，结合蛋白通过氢键结合到饱和非还原性末端残基的C4羟基在亚位点1。主要容纳饱和寡糖的M残基。在亚位点1，因为Ser-273与残基的羧基严格结合。在不饱和三糖（Δ GGG或Δ MMM）结合的AlgQ 1中，蛋白质与亚位点2和3处的底物羟基适当相互作用以容纳M或G，而底物羧基严格由亚位点2处的特定残基Tyr-129和亚位点3处的Lys-22识别。由于这种底物识别机制，菌株A1的溶质结合蛋白可以结合不同M/G比的杂多糖藻酸盐。
Alginate is a heteropolysaccharide that consists of beta-D-mannuronate (M) and alpha-L-guluronate (G). The Gram-negative bacterium Sphingomonas sp. A1 directly incorporates alginate into the cytoplasm through the periplasmic. solute-binding protein (AlgQ1 and AlgQ2)-dependent ABC transporter (AlgM1-AlgM2/AlgS-AlgS). Two binding proteins with at least four subsites strongly recognize the nonreducing terminal residue of alginate at subsite 1. Here, we show the broad substrate preference of strain A1 solute-binding proteins for M and G present in alginate and demonstrate the structural determinants in binding proteins for heteropolysaccharide recognition through X-ray crystallography of four AlgQ1 structures in complex with saturated and unsaturated alginate oligosaccharides. Alginates with different M/G ratios were assimilated by strain A1 cells and bound to AlgQ1 and AlgQ2. Crystal structures Of oligosaccharide-bound forms revealed that in addition to interaction between AlgQ1 and unsaturated oligosaccharides, the binding protein binds through hydrogen bonds to the C4 hydroxyl group of the saturated nonreducing terminal residue at subsite 1. The M residue of saturated oligosaccharides is predominantly accommodated. at subsite 1 because of the strict binding of Ser-273 to the carboxyl of the residue. In unsaturated trisaccharide (Delta GGG or Delta MMM)-bound AlgQ1, the protein, interacts appropriately with substrate hydroxyl groups at subsites 2 and 3 to accommodate M or G, while substrate carboxyl groups are strictly recognized by the specific residues Tyr-129 at subsite 2 and Lys-22 at subsite 3. Because of this substrate recognition mechanism, strain A1 solute,binding proteins can bind heteropolysaccharide alginate with different M/G ratios.