ISSN:
1432-072X
Keywords:
Key words Protein secretion
;
kil Gene
;
β-Glucanase
;
Escherichia coli
;
Growth-phase-dependent promoter
;
Stationary phase
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract Heterologous gene products produced by Escherichia coli cells can be exported into the culture medium by the action of the kil gene of the ColE1 plasmid, which encodes a bacterial release protein. The kil gene was fused with the stationary-phase promoter of the fic gene of E. coli, and a secretion cassette (Kil-Km cassette) containing the regulated kil gene, the Km-resistance gene, and multiple cloning sites for the integration of target genes was constructed. Using the gene for β-glucanase (bgl) as a target gene, it was shown that the protein produced was only secreted into the medium during the stationary phase. Quasi-lysis and lethality were not observed. The primary effect of the induction of the kil gene was the overproduction of β-glucanase. The total amount produced per milliliter of bacterial culture was almost threefold higher than that of the corresponding Kil– control. The protein pattern of periplasm and culture medium was analyzed before and after induction of the kil gene expression, indicating that the release of periplasmic proteins is semiselective. This secretion system is the first to use a growth-phase-regulated promoter for the expression of the kil gene.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/s002030050427
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