Non-native alpha-helices in the initial folding intermediate facilitate the ordered assembly of the beta-barrel in beta-lactoglobulin
Non-native alpha-helices in the initial folding intermediate facilitate the ordered assembly of the beta-barrel in beta-lactoglobulin
复制标题
初始折叠中间体中的非天然 α 螺旋促进 β 乳球蛋白中 β 桶的有序组装
DOI：
10.1021/acs.biochem.7b00458
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发表时间：
2017
期刊：
影响因子：
2.9
通讯作者：
Y.
中科院分区：
文献类型：
--
作者：
Sakurai;K.;Yagi;M.;Konuma;T.;Takahashi;S.;Nishimura;C.;Goto;Y.
多年来，人们已经研究了在β折叠蛋白的初始折叠阶段经常观察到的非天然α螺旋的作用。本文采用pH脉冲猝灭流标记法研究了牛β-乳球蛋白（β-lactoglobulin，βLG）几种突变体的残基水平结构。βLG在折叠的早期阶段呈现出一种具有非天然α螺旋结构（I 0）的塌陷中间体，尽管其天然形式主要由β结构组成。假野生型（WT*）βLG在I 0中的保护作用偏离了“折叠时平均掩埋面积”（AABUF）的模式。特别是，与AABUF相比，A链区域（非天然α-螺旋以I 0状态形成）的保护水平显著较低。G17 E，具有增加的螺旋倾向的突变体，显示出类似的保护模式。相反，E44 L（具有增加的β折叠倾向的突变体）对I 0的保护模式与WT* 不同，并且类似于AABUF模式。横向弛豫测量表明，在这些突变体的未折叠状态的残留结构的位置是一致的，在各自的I 0状态的保护残基。基于E44 L中I 0向天然状态的转化比WT* 慢，非天然α-螺旋通过阻止捕获折叠的相互作用促进β-桶的有序组装。
The roles of non-native α-helices frequently observed in the initial folding stage of β-sheet proteins have been examined for many years. We herein investigated the residue-level structures of several mutants of bovine β-lactoglobulin (βLG) in quenched-flow pH-pulse labeling experiments. βLG assumes a collapsed intermediate with a non-native α-helical structure (I0) in the early stage of folding, although its native form is predominantly composed of β-structures. The protection profile in I0of pseudo-wild type (WT*) βLG was found to deviate from the pattern of the “average area buried upon folding” (AABUF). In particular, the level of protection at the region of strand A, at which non-native α-helices form in the I0state, was significantly low compared to AABUF. G17E, the mutant with an increased helical propensity, showed a similar protection pattern. In contrast, the protection pattern for I0of E44L, the mutant with an increased β-sheet propensity, was distinct from that of WT* and resembled the AABUF pattern. Transverse relaxation measurements demonstrated that the positions of the residual structures in the unfolded states of these mutants were consistent with those of the protected residues in the respective I0states. On the basis of the slower conversion of I0to the native state for E44L to that for WT*, non-native α-helices facilitate the ordered assembly of the β-barrel by preventing interactions that trap folding.