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首页> 外文期刊>Journal of Molecular Biology >Transmembrane helix tilting and ligand-induced conformational changes in the lactose permease determined by site-directed chemical crosslinking in situ
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Transmembrane helix tilting and ligand-induced conformational changes in the lactose permease determined by site-directed chemical crosslinking in situ

机译：跨膜螺旋倾斜和配体诱导的乳糖通透酶构象变化通过定点化学原位交联测定

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The N-terminal six transmenbrane helices (N-6) and the C-terminal six transmembrane helices (C-6) of the lactose permease of Escherichia coli, each with a Cys residue, were co-expressed independently, and cross-linking was studied. Proximity of paired Cys residues in helices II (position 49, 52, 53, 56, 57, 60, 63 or 67) and VII (position 227, 230, 231, 234, 238, 241, 242 or 245) or XI (position 350, 353, 354, 357, 361 or 364) was examined by using two homobifunctional thiol-specific crosslinking agents of different lengths (6 or 10 Angstrom). The results demonstrate that a Cys residue placed in the periplasmic half of helix II (position 49, 52, 53 or 57) crosslinks to Cys residues in the periplasmic half of helix VII (position 241, 242 or 245). In contrast, no crosslinking is evident with paired-Cys residues in the cytoplasmic halves of helices LT (position 60, 63 or 67) and VII (position 227, 230, 231, 234 or 238). Remarkably, a Cys residue in the cytoplasmic half of helix II (position 60, 63 or 67) crosslinks with a Cys residue in the cytoplasmic half of helix XI (position 350, 353 or 354), while paired-Cys residues at positions in the periplasmic halves of the two helices do not crosslink. Therefore, helix II is tilted in such a manner that the periplasmic end is close to helix VII, and the cytoplasmic end is close to helix XI. Furthermore, ligand-binding alters the cross-linking efficiency of paired-Cys residues in helices II and VII or XI, indicating that both interfaces are conformationally active. The results are consistent with the conclusion that ligand-binding induces a scissors-like movement of helices II and VII that increases interhelical distance by 3 to 4 Angstrom at the periplasmic ends and decreases the distance by 3 to 4 Angstrom at the approximate middle of the two transmembrane helices. (C) 1998 Academic Press. [References: 44]
机译：大肠杆菌的乳糖通透酶的N末端的六个跨膜螺旋（N-6）和C末端的六个跨膜螺旋（C-6）各自共表达Cys残基，并且交联是研究。螺旋II（位置49、52、53、56、57、60、63或67）和VII（位置227、230、231、234、238、241、242或245）或XI（位置螺旋）中成对的Cys残基的邻近度通过使用两种不同长度（6或10埃）的同双功能硫醇特异性交联剂来检查（350、353、354、357、361或364）。结果表明，位于螺旋II的周质一半（位置49、52、53或57）中的Cys残基与螺旋VII的周质一半（位置241、242或245）中的Cys残基交联。相反，在螺旋LT（60、63或67位）和VII（227、230、231、234或238位）的胞质半部分中成对的Cys残基没有交联是明显的。值得注意的是，螺旋II的胞质半部分（位置60、63或67）中的Cys残基与螺旋XI的胞质半部分（位置350、353或354）中的Cys残基交联，而在螺旋II的位置中成对的Cys残基成对。两个螺旋的半周质不交联。因此，螺旋II以这样的方式倾斜：周质末端接近螺旋VII，而细胞质末端接近螺旋XI。此外，配体结合改变了螺旋II和VII或XI中成对的Cys残基的交联效率，表明这两个界面都在构象上具有活性。该结果与以下结论一致：配体结合诱导了螺旋状的II型和VII型剪刀状运动，该运动在周质末端使螺旋间距离增加了3至4埃，而在螺旋的大约中间使距离减少了3至4埃。两个跨膜螺旋。 （C）1998年学术出版社。 [参考：44]

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