Department of Life Sciences and Systems Biology, University of Torino, via Accademia Albertina 13, Torino 10123, Italy.
Biomolecules. 2013 Oct 8;3(4):778-811. doi: 10.3390/biom3040778.
Laboratory evolution techniques are becoming increasingly widespread among protein engineers for the development of novel and designed biocatalysts. The palette of different approaches ranges from complete randomized strategies to rational and structure-guided mutagenesis, with a wide variety of costs, impacts, drawbacks and relevance to biotechnology. A technique that convincingly compromises the extremes of fully randomized vs. rational mutagenesis, with a high benefit/cost ratio, is saturation mutagenesis. Here we will present and discuss this approach in its many facets, also tackling the issue of randomization, statistical evaluation of library completeness and throughput efficiency of screening methods. Successful recent applications covering different classes of enzymes will be presented referring to the literature and to research lines pursued in our group. The focus is put on saturation mutagenesis as a tool for designing novel biocatalysts specifically relevant to production of fine chemicals for improving bulk enzymes for industry and engineering technical enzymes involved in treatment of waste, detoxification and production of clean energy from renewable sources.
实验室进化技术在蛋白质工程师中越来越普及，用于开发新型和设计的生物催化剂。不同方法的范围从完全随机的策略到理性和结构导向的诱变，具有广泛的成本、影响、缺点和与生物技术的相关性。一种技术有效地平衡了完全随机与理性诱变的极端情况，具有高的收益/成本比，这就是饱和突变。在这里，我们将介绍和讨论这种方法的许多方面，还将讨论随机化问题、文库完整性的统计评估以及筛选方法的通量效率。我们将参考文献和我们小组的研究方向，介绍最近在不同酶类方面的成功应用。重点是将饱和突变作为一种工具，用于设计新型生物催化剂，特别是针对精细化学品生产的生物催化剂，以改进工业用大宗酶和用于处理废物、解毒和从可再生资源生产清洁能源的工程技术酶。
