Synthesis of human renalase1 in Escherichia coli and its purification as a FAD-containing holoprotein.
Protein Expr Purif
; 72(2): 244-53, 2010 Aug.
Article
em En
| MEDLINE
| ID: mdl-20302943
ABSTRACT
Renalase is a protein ubiquitous in vertebrates, which has been proposed to modulate blood pressure and heart rate, and whose downregulation might result in hypertension. Despite its potential relevance for human health, the biochemical characterization of renalase is still lacking, possibly due to difficulties in obtaining it in recombinant form. By expressing two different gene constructs, we found that the major isoform of human renalase, renalase1, is mainly produced in Escherichia coli in inclusion bodies. However, by optimizing the expression conditions, significant amounts of soluble products were obtained. Both soluble renalase forms have been purified to homogeneity exploiting their N-terminal His-tag. Linking of the protein of interest to the SUMO protein did not improve solubility, but yielded untagged renalase1 after proteolytic processing of the fusion product. The two recombinant renalase forms displayed the same molecular properties. They bind equimolar amounts of FAD and appear to be correctly folded by various criteria. The procedures for the production and isolation of recombinant renalase1 here reported are expected to boost the much awaited biochemical studies on this remarkable protein.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas Recombinantes
/
Flavina-Adenina Dinucleotídeo
/
Monoaminoxidase
Limite:
Animals
/
Humans
/
Male
Idioma:
En
Revista:
Protein Expr Purif
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2010
Tipo de documento:
Article
País de afiliação:
Itália