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首页> 外文期刊>Biophysical Chemistry: An International Journal Devoted to the Physical Chemistry of Biological Phenomena >A molecular dynamics model of the Bt toxin Cyt1A and its validation by resonance energy transfer.
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A molecular dynamics model of the Bt toxin Cyt1A and its validation by resonance energy transfer.

机译：Bt毒素Cyt1A的分子动力学模型及其通过共振能量转移的验证。

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摘要

Cyt1A is a cytolytic toxin from Bacillus thuringiensis var. israelensis. A computer model of the toxin in solution was generated and validated by resonance energy transfer (RET). The average distance between the two tryptophans (residues 158 and 161) and the fluorescently labeled cysteine 190 was 2.16 nm, which closely matched the distance predicted in computer simulations, 2.2 nm. The simulation results were able to explain two previous experimental observations: (i) amino-acid sequences of all Cyt toxins contain four blocks of highly conserved residues; and (ii) several single-point mutations drastically abrogated Cyt1A's toxicity. Selective randomization of atomic coordinates in the computer model revealed that the conserved blocks are important for proper folding and stability of the toxin molecule. Replacing lysine 225 with alanine, a mutation that renders the toxin inactive, was shown to result in breaking the hydrogen bonds between K225 and V126, L123, and Y189. Calculated Helmholtz free energy difference of the inactive mutation K225A was higher by 12 kcal/mol and 5 kcal/mol than the values for the benign mutations K118A and K198A, respectively, which indicates that the K225A mutant is significantly destabilized. The normal-mode and principal-component analyses revealed that in the wild-type Cyt1A the region around the residue K225 is quite stationary, due to the hydrogen-bond network around K225. In contrast, pronounced twisting and stretching were observed in the mutant K225A, and the region around the residue K225 becomes unstable. Our results indicate that conformational differences in this mutant spread far away from the site of the mutation, suggesting that the mutant is inactivated due to an overall change in conformation and diminished stability rather than due to a localized alteration of a "binding" or "active" site.
机译：Cyt1A是苏云金芽孢杆菌变种的溶细胞毒素。以色列。生成了溶液中毒素的计算机模型，并通过共振能量转移（RET）进行了验证。两个色氨酸（残基158和161）与荧光标记的半胱氨酸190之间的平均距离为2.16 nm，与计算机模拟中预测的2.2 nm紧密匹配。模拟结果能够解释先前的两个实验观察结果：（i）所有Cyt毒素的氨基酸序列均包含四个高度保守的残基嵌段; （ii）多个单点突变彻底废除了Cyt1A的毒性。在计算机模型中原子坐标的选择性随机化表明，保守的嵌段对于毒素分子的正确折叠和稳定性很重要。用丙氨酸代替赖氨酸225可以使K225和V126，L123和Y189之间的氢键断裂，该突变使毒素失去活性。失活突变K225A的计算的亥姆霍兹自由能差分别比良性突变K118A和K198A的值高12 kcal / mol和5 kcal / mol，这表明K225A突变体明显不稳定。正常模式和主成分分析显示，在野生型Cyt1A中，由于K225周围的氢键网络，残基K225周围的区域非常稳定。相反，在突变体K225A中观察到明显的扭曲和拉伸，并且残基K225周围的区域变得不稳定。我们的结果表明该突变体的构象差异远离突变位点扩散，这表明该突变体因构象的整体变化和稳定性降低而失活，而不是由于“结合”或“活性”的局部改变”网站。

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