Title:High Throughput Screening of Esterases, Lipases and Phospholipases in Mutant and Metagenomic Libraries: A Review
VOLUME: 19 ISSUE: 8
Author(s):Carlina Peña-García, Mónica Martínez-Martínez, Dolores Reyes-Duarte* and Manuel Ferrer*
Affiliation:Departamento de Procesos y Tecnología, Universidad Autónoma Metropolitana, Unidad Cuajimalpa, Av. Vasco de Quiroga 4871, Col. Santa Fe, Deleg, Cuajimalpa, 05348, CDMX, México., Institute of Catalysis, Consejo Superior de Investigaciones Científicas (CSIC), Marie Curie 2, 28049 Madrid, Spain.
Keywords:Biocatalysts, esterases, high throughput screening, lipases, metagenomic, phospholipases, protein engineering.
Abstract:Nowadays, enzymes can be efficiently identified and screened from
metagenomic resources or mutant libraries. A set of a few hundred new enzymes can
be found using a simple substrate within few months. Hence, the establishment of
collections of enzymes is no longer a big hurdle. However, a key problem is the
relatively low rate of positive hits and that a timeline of several years from the
identification of a gene to the development of a process is the reality rather than the
exception. Major problems are related to the time-consuming and cost-intensive
screening process that only very few enzymes finally pass. Accessing to the highest
possible enzyme and mutant diversity by different, but complementary approaches is
increasingly important. The aim of this review is to deliver state-of-art status of
traditional and novel screening protocols for targeting lipases, esterases and phospholipases of
industrial relevance, and that can be applied at high throughput scale (HTS) for at least 200 distinct
substrates, at a speed of more than 105 – 108 clones/day. We also review fine-tuning sequence analysis
pipelines and in silico tools, which can further improve enzyme selection by an unprecedent speed (up
to 1030 enzymes). If the hit rate in an enzyme collection could be increased by HTS approaches, it can
be expected that also the very further expensive and time-consuming enzyme optimization phase could
be significantly shortened, as the processes of enzyme-candidate selection by such methods can be
adapted to conditions most likely similar to the ones needed at industrial scale.
