A novel reporter system monitoring sortase A catalyzed protein ligation efficiency / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 284-293, 2014.
Artigo
em Chinês
| WPRIM
| ID: wpr-279522
ABSTRACT
Efforts on directed evolution of sortase A to optimize its catalytic properties have been undertaken and shown the promise. To facilitate screening of sortase A mutants with expected catalytic properties, a novel ligation efficiency monitoring system, including reporter substrates GFP-LPETG and GGGYK-Biotin, was developed. GFP-LPETG, wild type sortase A, and a recently reported high activity sortase A mutant were prepared recombinantly from Escherichia coli BL21 (DE3). Taking advantage of the newly designed reporter system, the ligation efficiency catalyzed by wild type and mutant form of sortase A could be sensitively monitored via SDS-PAGE directly. Consistent with previous report, the mutant sortase A displayed much higher catalytic activity compared to wild type enzyme, indicating the new reporter system is easily and fast handled and sensitive. The application of this reporter system into systemic screening will facilitate future directed optimization of sortase A.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Proteínas de Bactérias
/
Biotina
/
Cisteína Endopeptidases
/
Genes Reporter
/
Aminoaciltransferases
/
Eletroforese em Gel de Poliacrilamida
/
Escherichia coli
/
Proteínas Mutantes
/
Biocatálise
/
Genética
Idioma:
Chinês
Revista:
Chinese Journal of Biotechnology
Ano de publicação:
2014
Tipo de documento:
Artigo
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