Cloning-Independent Expression and Analysis of ω-Transaminases by Use of a Cell-Free Protein Synthesis System
2010; American Society for Microbiology; Volume: 76; Issue: 18 Linguagem: Inglês
10.1128/aem.00029-10
ISSN1098-5336
AutoresYong‐Chan Kwon, Kyung‐Ho Lee, Ho‐Cheol Kim, Kyuboem Han, Joo‐Hyun Seo, Byung‐Gee Kim, Dong‐Myung Kim,
Tópico(s)Biochemical and Molecular Research
ResumoABSTRACT Herewith we report the expression and screening of microbial enzymes without involving cloning procedures. Computationally predicted putative ω-transaminase (ω-TA) genes were PCR amplified from the bacterial colonies and expressed in a cell-free protein synthesis system for subsequent analysis of their enzymatic activity and substrate specificity. Through the cell-free expression analysis of the putative ω-TA genes, a number of enzyme-substrate pairs were identified in a matter of hours. We expect that the proposed strategy will provide a universal platform for bridging the information gap between nucleotide sequence and protein function to accelerate the discovery of novel enzymes.
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