Functioning of a hybrid BRP-beta-lactamase protein in the release of cloacin DF13 and lysis of Escherichia coli cells.
The gene encoding a hybrid BRP-Bla protein consisting of the pCloDF13 encoded BRP signal sequence, 25 of the 28 amino acid residues of the mature bacteriocin release protein (BRP) and the mature portion of beta-lactamase (Bla) was subcloned in the expression vector pEB112. A similar construct was made using a mutant gene encoding a BRP-Bla protein in which the cysteine residue at the +1 position was changed into a glycine residue. The expression, processing, functioning and subcellular localization of the 'wild-type' and mutant hybrid protein at high-level expression conditions were studied. The 'wild-type' BRP-Bla protein was mainly found in the outer membranes and possessed all the activities of the BRP itself; the protein was able to bring about the release of cloacin DF13 and caused apparent cell-lysis after high-level synthesis. The mutant hybrid protein was predominantly located in the inner membranes, was inactive in the release of cloacin DF13, but caused apparent cell-lysis only after strong induction.
Luirink
J
J
Department of Molecular Microbiology, Vrije Universiteit, Amsterdam, The Netherlands.
de Jong
J
J
van Putten
A J
AJ
de Graaf
F K
FK
Oudega
B
B
D001425:Bacterial Outer Membrane Proteins / Q000032:analysis; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D001430:Bacteriocins / Q000235:genetics*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000254:growth & development; D029968:Escherichia coli Proteins; D006824:Hybridization, Genetic; D009154:Mutation; D001618:beta-Lactamases / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics*
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D001430:Bacteriocins; D029968:Escherichia coli Proteins; C052368:cloacin DF13-encoded bacteriocin release protein, E coli; D001618:beta-Lactamases