The location of the ligand-binding site of carbohydrate-binding modules that have evolved from a common sequence is not conserved
The location of the ligand-binding site of carbohydrate-binding modules that have evolved from a common sequence is not conserved
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DOI：
10.1074/jbc.m109142200
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发表时间：
2001-12-21
影响因子：
4.8
通讯作者：
Gilbert, HJ
中科院分区：
文献类型：
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作者：
Czjzek, M;Bolam, DN;Gilbert, HJ
多糖降解酶通常是含有非催化碳水化合物结合模块（CBM）的模块蛋白，其增强催化模块的活性。建立信任措施已被分为基于序列的家庭，和三维结构数据可用于这些家庭的一半。Clostridium thermocellum木聚糖酶11 A是一种模块化酶，其含有来自家族6（CBM 6）的CBM，其结构数据不可用。我们已经确定了该模块的晶体结构，分辨率为2.1埃。该蛋白是一种β-夹心，含有两个潜在的配体结合裂缝，命名为裂缝A和B。CBM主要与木聚糖相互作用，并且NMR光谱加上定点诱变鉴定了含有Trp-92、Tyr-34和Asn-120的裂缝A作为配体结合位点。CBM 6的总体折叠与CBM家族4和22中的蛋白质相似，尽管令人惊讶地，CBM 4和CBM 22中的配体结合位点与CBM 6中的裂缝B相当。这些结构数据定义了CBM超家族，包括CBM 4、CBM 6和CBM 22，并表明，尽管CBM是从相对较少的祖先进化而来的，但参与配体识别的结构元件已在不同位置组装在祖先支架上。
Polysaccharide-degrading enzymes are generally modular proteins that contain non-catalytic carbohydrate-binding modules (CBMs), which potentiate the activity of the catalytic module. CBMs have been grouped into sequence-based families, and three-dimensional structural data are available for half of these families. Clostridium thermocellum xylanase 11A is a modular enzyme that contains a CBM from family 6 (CBM6), for which no structural data are available. We have determined the crystal structure of this module to a resolution of 2.1 Angstrom. The protein is a beta -sandwich that contains two potential ligand-binding clefts designated cleft A and B. The CBM interacts primarily with xylan, and NMR spectroscopy coupled with site-directed mutagenesis identified cleft A, containing Trp-92, Tyr-34, and Asn-120, as the ligand-binding site. The overall fold of CBM6 is similar to proteins in CBM families 4 and 22, although surprisingly the ligand-binding site in CBM4 and CBM22 is equivalent to cleft B in CBM6. These structural data define a superfamily of CBMs, comprising CBM4, CBM6, and CBM22, and demonstrate that, although CBMs have evolved from a relatively small number of ancestors, the structural elements involved in ligand recognition have been assembled at different locations on the ancestral scaffold.