Extraction of recombinant protein from Escherichia coli by using a novel cell autolysis activity of VanX.
Anal Biochem
; 439(2): 212-7, 2013 Aug 15.
Article
en En
| MEDLINE
| ID: mdl-23624113
ABSTRACT
Escherichia coli is a versatile, low-cost, and popular host for expressing recombinant proteins. However, extracting recombinant proteins from E. coli requires cell wall breakage, which is both time- and effort-consuming. Here we report a novel cell breakage method based on our recent finding that VanX, which is a d-Ala-d-Ala dipeptidase encoded in a vancomycin-resistant VanA gene cluster, exhibits a strong cell lysis activity when expressed in isolation in E. coli. In our strategy, we coexpress VanX with the target protein, causing cell autolysis and release of the cellular content into the culture medium. We demonstrated this strategy for two model proteins, a green fluorescent protein variant (GFPuv) and Gaussia luciferase, and optimized the autolysis conditions and coexpression vectors. The fluorescence activity of GFPuv collected from the medium was identical to that of GFPuv purified by conventional methods. Cell breakage by VanX-mediated autolysis is very simple to implement and will efficiently complement traditional methods.
Palabras clave
Texto completo:
1
Banco de datos:
MEDLINE
Asunto principal:
Proteínas Bacterianas
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Bacteriólisis
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D-Ala-D-Ala Carboxipeptidasa de Tipo Serina
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Proteínas Fluorescentes Verdes
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Escherichia coli
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Luciferasas
Tipo de estudio:
Prognostic_studies
Idioma:
En
Año:
2013
Tipo del documento:
Article