Regulated recombinant protein production in the Antarctic bacterium Pseudoalteromonas haloplanktis TAC125.
Methods Mol Biol
; 824: 203-18, 2012.
Article
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| MEDLINE
| ID: mdl-22160900
ABSTRACT
This review reports results from our laboratory on the development of an effective inducible expression system for the homologous/heterologous protein production in cold-adapted bacteria. Recently, we isolated and characterized a regulative genomic region from Pseudoalteromonas haloplanktis TAC125; in particular, a two-component regulatory system was identified. It is involved in the transcriptional regulation of the gene coding for an outer membrane porin (PSHAb0363) that is strongly induced by the presence of L -malate in the growth medium.We used the regulative region comprising the two-component system located upstream the PSHAb0363 gene to construct an inducible expression vector - named pUCRP - under the control of L -malate. Performances of the inducible system were tested using the psychrophilic ß-galactosidase from P. haloplanktis TAE79 as model enzyme to be produced. Our results demonstrate that the recombinant cold-adapted enzyme is produced in P. haloplanktis TAC125 in good yields and in a completely soluble and catalytically competent form. Moreover, an evaluation of optimal induction conditions for protein production was also carried out in two consecutive steps:
(1) definition of the optimal cellular growth phase in which the gene expression has to be induced; (2) definition of the optimal inducer concentration that has to be added in the growth medium.
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
Proteínas Recombinantes
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Adaptación Biológica
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Regulación Bacteriana de la Expresión Génica
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Frío
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Pseudoalteromonas
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2012
Tipo del documento:
Article
País de afiliación:
Italia