Functional annotation of the mesophilic-like character of mutants in a cold-adapted enzyme by self-organising map analysis of their molecular dynamics
Functional annotation of the mesophilic-like character of mutants in a cold-adapted enzyme by self-organising map analysis of their molecular dynamics
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DOI：
10.1039/c2mb25192b
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发表时间：
2012-01-01
影响因子：
--
通讯作者：
Papaleo, Elena
中科院分区：
文献类型：
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作者：
Fraccalvieri, Domenico;Tiberti, Matteo;Papaleo, Elena
多重比较的分子动力学（MD）轨迹的突变体在冷适应的α-淀粉酶（AHA）可以用来阐明恢复嗜温样活性所需的功能特征。不幸的是，它是具有挑战性的，以确定不同的动态行为，并正确地将它们与功能活动的常规分析。我们在这里采用了一个以前开发的和强大的两阶段的方法，结合自组织地图（SOM）和层次聚类比较蛋白质的构象合奏。此外，我们设计了一种新的策略，以确定特定的突变，更有效地转换的嗜冷酶（AHA）的动态签名的嗜温对应物（PPA）。在AHA及其变体上训练的SOM用于对PPA MD系综进行分类，并成功地突出了目标酶的灵活性和不同突变体之间的关系。此外，突变体的局部特征，主要影响他们的整体灵活性在一个mesophilic-like方向被检测到。事实证明，突变的冷适应酶的疏水性和芳香族残基是最有效的恢复PPA的动态功能，并可以指导更多的嗜温突变体的设计。总之，我们的策略可以有效地提取特定的动态签名相关的功能从多重比较的MD构象合奏。因此，它可以是一个有前途的工具，蛋白质工程。
Multiple comparison of the Molecular Dynamics (MD) trajectories of mutants in a cold-adapted alpha-amylase (AHA) could be used to elucidate functional features required to restore mesophilic-like activity. Unfortunately it is challenging to identify the different dynamic behaviors and correctly relate them to functional activity by routine analysis. We here employed a previously developed and robust two-stage approach that combines Self-Organising Maps (SOMs) and hierarchical clustering to compare conformational ensembles of proteins. Moreover, we designed a novel strategy to identify the specific mutations that more efficiently convert the dynamic signature of the psychrophilic enzyme (AHA) to that of the mesophilic counterpart (PPA). The SOM trained on AHA and its variants was used to classify a PPA MD ensemble and successfully highlighted the relationships between the flexibilities of the target enzyme and of the different mutants. Moreover the local features of the mutants that mostly influence their global flexibility in a mesophilic-like direction were detected. It turns out that mutations of the cold-adapted enzyme to hydrophobic and aromatic residues are the most effective in restoring the PPA dynamic features and could guide the design of more mesophilic-like mutants. In conclusion, our strategy can efficiently extract specific dynamic signatures related to function from multiple comparisons of MD conformational ensembles. Therefore, it can be a promising tool for protein engineering.