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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Dehydration of main-chain amides in the final folding step of single-chain monellin revealed by time-resolved infrared spectroscopy
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Dehydration of main-chain amides in the final folding step of single-chain monellin revealed by time-resolved infrared spectroscopy

机译：时间分辨红外光谱显示单链莫内林最后折叠步骤中主链酰胺的脱水

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摘要

Kinetic IR spectroscopy was used to reveal β-sheet formation and water expulsion in the folding of single-chain monellin (SMN) composed of a five-stranded β-sheet and an α-helix. The time-resolved IR spectra between 100 μs and 10 s were analyzed based on two consecutive intermediates, I_1 and I_2, appearing within 100 μs and with a time constant of ≈100 ms, respectively. The initial unfolded state showed broad amide I' corresponded to a fluctuating conformation. In contrast, I_1 possessed a feature at 1,636 cm~(-1) for solvated helix and weak features assignable to turns, demonstrating the rapid formation of helix and turns. I_2 possessed a line for solvated helix at 1,637 cm~(-1) and major and minor lines for β-sheet at 1,625 and 1,680 cm~(-1), respectively. The splitting of the major and minor lines is smaller than that of the native state, implying an incomplete formation of the β-sheet. Furthermore, both major and minor lines demonstrated a low-frequency shift compared to those of the native state, which was interpreted to be caused by hydration of the C=O group in the β-sheet. Together with the identification of solvated helix, the core domain of I_2 was interpreted as being hydrated. Finally, slow conversion of the water-penetrated core of I_2 to the dehydrated core of the native state was observed. We propose that both the expulsion of water, hydrogen-bonded to main-chain amides, and the completion of the secondary structure formation contribute to the energetic barrier of the rate-limiting step in SMN folding.
机译：动力学红外光谱用于揭示由五链β-折叠和α-螺旋组成的单链莫奈林（SMN）折叠时β-折叠的形成和水的排出。基于两个连续的中间体I_1和I_2，分析了100μs和10 s之间的时间分辨IR光谱，它们分别出现在100μs内并且具有约100 ms的时间常数。最初的展开状态显示宽酰胺I'对应于波动的构象。相反，I_1在1636 cm〜（-1）处具有溶剂化螺旋的特征，而较弱的特征可分配给转弯，表明螺旋和转弯的快速形成。 I_2在1637 cm〜（-1）处有一条溶剂化螺旋线，在β-折叠处有1,625 cm〜（-1）的主线和副线。主线和次线的分裂小于原始状态的分裂，这意味着β-折叠的形成不完整。此外，与原始状态相比，主线和次线均表现出低频偏移，这被认为是由β-折叠中C = O基团的水合引起的。与溶剂化螺旋的鉴定一起，I_2的核心结构域被解释为是水合的。最后，观察到I_2的透水核心缓慢转化为天然状态的脱水核心。我们提出，水的排出，氢键合到主链酰胺上，以及二级结构的形成都有助于SMN折叠中限速步骤的能量屏障。

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