Sugar binding induces the same global conformational change in purified LacY as in the native bacterial membrane
Sugar binding induces the same global conformational change in purified LacY as in the native bacterial membrane
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DOI：
10.1073/pnas.1004515107
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发表时间：
2010-05-25
影响因子：
11.1
通讯作者：
Kaback, H. Ronald
中科院分区：
文献类型：
--
作者：
Nie, Yiling;Kaback, H. Ronald
许多独立的证据表明，大肠杆菌（LacY）的乳糖通透酶是高度动态的，糖结合导致关闭一个大的面向内的空腔与开放的一个广泛的面向外的亲水性空腔。因此，乳糖/H+同向转运催化的LacY很可能涉及一个全球性的构象变化，允许交替访问单一的糖和H+结合位点的膜的两侧（交替访问模型）。LacY的X射线晶体结构以及大多数光谱研究都使用洗涤剂胶束中的纯化蛋白质。通过使用定点烷基化，我们现在证明，糖结合诱导几乎相同的全球构象变化的LacY蛋白质是否在天然细菌膜或溶解和纯化的洗涤剂。结果还表明，X射线晶体结构反映了在不存在糖的情况下天然膜中野生型LacY的结构。
Many independent lines of evidence indicate that the lactose permease of Escherichia coli (LacY) is highly dynamic and that sugar binding causes closing of a large inward-facing cavity with opening of a wide outward-facing hydrophilic cavity. Therefore, lactose/H+ symport catalyzed by LacY very likely involves a global conformational change that allows alternating access of single sugar- and H+-binding sites to either side of the membrane (the alternating access model). The x-ray crystal structures of LacY, as well as the majority of spectroscopic studies, use purified protein in detergent micelles. By using site-directed alkylation, we now demonstrate that sugar binding induces virtually the same global conformational change in LacY whether the protein is in the native bacterial membrane or is solubilized and purified in detergent. The results also indicate that the x-ray crystal structure reflects the structure of wild-type LacY in the native membrane in the absence of sugar.