Nakano Shogo, Asano Yasuhisa
1] Biotechnology Research Center and Department of Biotechnology, Toyama Prefectural University, 5180 Kurokawa, Imizu, Toyama 939-0398, Japan [2] Asano Active Enzyme Molecule Project, ERATO, JST, 5180 Kurokawa, Imizu, Toyama 939-0398, Japan.
Sci Rep. 2015 Feb 3;5:8193. doi: 10.1038/srep08193.
Development of software and methods for design of complete sequences of functional proteins could contribute to studies of protein engineering and protein evolution. To this end, we developed the INTMSAlign software, and used it to design functional proteins and evaluate their usefulness. The software could assign both consensus and correlation residues of target proteins. We generated three protein sequences with S-selective hydroxynitrile lyase (S-HNL) activity, which we call designed S-HNLs; these proteins folded as efficiently as the native S-HNL. Sequence and biochemical analysis of the designed S-HNLs suggested that accumulation of neutral mutations occurs during the process of S-HNLs evolution from a low-activity form to a high-activity (native) form. Taken together, our results demonstrate that our software and the associated methods could be applied not only to design of complete sequences, but also to predictions of protein evolution, especially within families such as esterases and S-HNLs.
用于设计功能蛋白完整序列的软件和方法的开发，可能有助于蛋白质工程和蛋白质进化的研究。为此，我们开发了INTMSAlign软件，并使用它来设计功能蛋白并评估其效用。该软件可以确定目标蛋白的共有残基和相关残基。我们生成了三个具有S-选择性羟腈裂解酶（S-HNL）活性的蛋白序列，我们将其称为设计型S-HNL；这些蛋白的折叠效率与天然S-HNL一样高。对设计型S-HNL的序列和生化分析表明，在S-HNL从低活性形式进化到高活性（天然）形式的过程中会发生中性突变的积累。综上所述，我们的结果表明，我们的软件和相关方法不仅可以应用于完整序列的设计，还可以用于蛋白质进化的预测，特别是在酯酶和S-HNL等家族中。
