Expression of cold-adapted ß-1,3-xylanase as a fusion protein with a ProS2 tag and purification using immobilized metal affinity chromatography with a high concentration of ArgHCl.
Biotechnol Lett
; 37(1): 89-94, 2015 Jan.
Article
en En
| MEDLINE
| ID: mdl-25214227
ABSTRACT
Cold-adapted ß-1,3-xylanase (P.t.Xyn26A) from the psychrotrophic bacterium, Psychroflexus torquis, was expressed as a fusion protein with tandem repeats of the N-terminal domain of Protein S from Myxocuccus xanthus (ProS2) in Escherichia coli. After cell lysis in phosphate buffer, most of the ProS2-P.t.Xyn26A was located in the insoluble fraction and aggregated during purification. Arginine hydrochloride (ArgHCl) efficiently solubilized the ProS2-P.t.Xyn26A. The solubilized ProS2-P.t.Xyn26A was purified using immobilized metal affinity chromatography (IMAC) with 500 mM ArgHCl. After cleavage of ProS2-P.t.Xyn26A by human rhinovirus 3C protease, we confirmed that recombinant P.t.Xyn26A maintained its native fold. This is the first report of the expression of a cold-adapted enzyme fused with a ProS2 tag under IMAC purification using a high concentration of ArgHCl. These insights into the expression and purification should be useful during the handling of cold-adapted enzymes.
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Bases de datos:
MEDLINE
Asunto principal:
Arginina
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Proteínas Bacterianas
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Proteínas Recombinantes de Fusión
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Cromatografía de Afinidad
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Xilano Endo-1,3-beta-Xilosidasa
Idioma:
En
Revista:
Biotechnol Lett
Año:
2015
Tipo del documento:
Article
País de afiliación:
Japón