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Enzyme and microbial technology

Establishment and application of a modified membrane-blot assay for Rhizomucor miehei lipases aimed at improving their methanol tolerance and thermostability.


PMID 28465058

Abstract

Directed evolution has been proved an effective way to improve the stability of proteins, but high throughput screening assays for directed evolution with simultaneous improvement of two or more properties are still rare. In this study, we aimed to establish a membrane-blot assay for use in the high-throughput screening of Rhizomucor miehei lipases (RMLs). With the assistance of the membrane-blot screening assay, a mutant E47K named G10 that showed improved thermal stability was detected in the first round of error-prone PCR. Using G10 as the parent, two variants G10-11 and G10-20 that showed improved thermal stability and methanol tolerance without loss of activity compared to the wild type RML were obtained. The T

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N7653
p-Nitrophenyl Phosphate Liquid Substrate System, liquid
C6H4NNa2O6P