Affiliation:
1. Department of Molecular Microbiology, Institute of Molecular Biological Sciences, BioCentrum Amsterdam Faculty of Biology, Vrije Universiteit, 1081 HV Amsterdam, The Netherlands
Abstract
ABSTRACT
Bacteriocin release proteins (BRPs) can be used for the release of heterologous proteins from the
Escherichia coli
periplasm into the culture medium. However, high-level expression of BRP causes apparent lysis of the host cells in liquid cultures (quasi-lysis) and inhibition of growth on broth agar plates (lethality). To optimize BRP-mediated protein release, the pCloDF13 BRP gene was subjected to random mutagenesis by using PCR techniques. Mutated BRPs with a strongly reduced capacity to cause growth inhibition on broth agar plates were selected, analyzed by nucleotide sequencing, and further characterized by performing growth and release experiments in liquid cultures. A subset of these BRP derivatives did not cause quasi-lysis and had only a small effect on growth but still functioned in the release of the periplasmic protein β-lactamase and the periplasmic K88 molecular chaperone FaeE and in the release of the bacteriocin cloacin DF13 into the culture medium. These BRP derivatives can be more efficiently used for extracellular production of proteins by
E. coli
than can the original BRP.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology
Cited by
12 articles.
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