Institute for Microbiology, Technische Universität Dresden, Dresden, Germany.
Enzyme Process Technology, RWTH Aachen University, Aachener Verfahrenstechnik, Aachen, Germany.
J Appl Microbiol. 2018 Feb;124(2):480-490. doi: 10.1111/jam.13664.
The purpose of the study was to demonstrate feasibility of the Conserved Domains Database (CDD) for identification of novel biocatalysts with desirable properties from a class of well-characterized biocatalysts.
The thermostable ADH from Sulfolobus solfataricus with a broad substrate range was applied as a template for the search for novel thermostable ADHs via CDD. From the resulting hits, a putative ADH gene from the thermophilic organism Chloroflexus aurantiacus was cloned and expressed in Escherichia coli. The resulting enzyme was purified and characterized. With a temperature activity optimum of 70°C and a broad substrate spectrum especially for diketones, a versatile new biocatalyst was obtained.
Database-based mining in CDD is a suitable approach to obtain novel biocatalysts with desirable properties. Thereby, the available diversity of similar but not equal enzymes within this class can be increased.
For industrial applications, there is a demand for larger diversity of similar well-characterized enzymes in order to test them for a given process (biodiversity screening). For fundamental science, the comparison of enzymes with similar function but different sequence can provide insight into structure function relationships or the evolution of enzymes. This study gives a good example on how this demand can be efficiently met.
本研究旨在展示保守结构域数据库（CDD）在从一类特征明确的生物催化剂中鉴定具有理想特性的新型生物催化剂方面的可行性。
应用具有广泛底物范围的嗜热菌 Sulfolobus solfataricus 的热稳定醇脱氢酶（ADH）作为模板，通过 CDD 搜索新型热稳定 ADH。从所得命中结果中，克隆并在大肠杆菌中表达了来自嗜热生物 Chloroflexus aurantiacus 的假定 ADH 基因。所得酶被纯化并进行了表征。该酶具有 70°C 的温度活性最优值和广泛的底物谱，特别是对二酮，是一种多功能的新型生物催化剂。
基于数据库的 CDD 挖掘是获得具有理想特性的新型生物催化剂的合适方法。因此，可以增加该类中类似但不相同的酶的可用多样性。
对于工业应用，需要更大的类似经过充分研究的酶的多样性，以便对给定的过程（生物多样性筛选）进行测试。对于基础科学，具有相似功能但不同序列的酶的比较可以深入了解结构功能关系或酶的进化。本研究很好地说明了如何有效地满足这一需求。
