Estimating the success of enzyme bioprospecting through metagenomics: current status and future trends.
Estimating the success of enzyme bioprospecting through metagenomics: current status and future trends.
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DOI：
10.1111/1751-7915.12309
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发表时间：
2016-01
影响因子：
5.7
通讯作者：
Golyshin PN
中科院分区：
文献类型：
--
作者：
Ferrer M;Martínez-Martínez M;Bargiela R;Streit WR;Golyshina OV;Golyshin PN
最近的报告表明，从环境基因组中建立工业相关的酶集合已成为一种常规程序。在评估的研究中，使用初始筛选方案在平均约53 000个克隆中获得平均约44个活性克隆。当选择通过直接测序获得的新型宏基因组酶序列并进行高通量表达用于后续生产和表征时，该数量可以在更短的时间内显著增加。通过初始筛选对克隆文库进行预筛选，然后对插入片段进行焦磷酸测序，使得鉴定编码感兴趣的酶的基因的成功率提高了106倍。然而，从酶鉴定到建立工业过程需要更长的时间，通常为数年。如果在真实的应用条件下以高周转率进行的酶鉴定的命中频率可以增加，同时仍然覆盖高天然多样性，则非常昂贵和耗时的酶优化阶段可能会显著缩短。在这一点上，重要的是要审查目前的知识，成功的微调天真和序列为基础的筛选协议的酶选择，并描述世界各地的环境，已经通过宏基因组学工具进行酶筛选计划。在这里，我们提供了这样的估计，并提出了当前的挑战和未来的行动之前，环境酶可以成功地引入市场。
Recent reports have suggested that the establishment of industrially relevant enzyme collections from environmental genomes has become a routine procedure. Across the studies assessed, a mean number of approximately 44 active clones were obtained in an average size of approximately 53 000 clones tested using naïve screening protocols. This number could be significantly increased in shorter times when novel metagenome enzyme sequences obtained by direct sequencing are selected and subjected to high‐throughput expression for subsequent production and characterization. The pre‐screening of clone libraries by naïve screens followed by the pyrosequencing of the inserts allowed for a 106‐fold increase in the success rate of identifying genes encoding enzymes of interest. However, a much longer time, usually on the order of years, is needed from the time of enzyme identification to the establishment of an industrial process. If the hit frequency for the identification of enzymes performing at high turnover rates under real application conditions could be increased while still covering a high natural diversity, the very expensive and time‐consuming enzyme optimization phase would likely be significantly shortened. At this point, it is important to review the current knowledge about the success of fine‐tuned naïve‐ and sequence‐based screening protocols for enzyme selection and to describe the environments worldwide that have already been subjected to enzyme screen programmes through metagenomic tools. Here, we provide such estimations and suggest the current challenges and future actions needed before environmental enzymes can be successfully introduced into the market.