Abstract
Many non-lantibiotic bacteriocins of lactic acid bacteria are produced as precursors with N-terminal leader peptides different from those present in preproteins exported by the general sec-dependent (type II) secretion pathway. These bacteriocins utilize a dedicated (type I) secretion system for externalization. The secretion apparatus for the lactococcins A, B, and M/N (LcnA, B, and M/N) from Lactococcus lactis is composed of the two membrane proteins LcnC and LcnD. LcnC belongs to the ATP-binding cassette transporters, whereas LcnD is a protein with similarities to other accessory proteins of type I secretion systems. This paper shows that the N-terminal part of LcnC is involved in the processing of the precursor of LcnA. By making translational fusions of LcnC to the reporter proteins beta-galactosidase (LacZ) and alkaline phosphatase (PhoA*), it was shown that both the N- and C-terminal parts of LcnC are located in the cytoplasm. As the N terminus of LcnC is required for LcnA maturation and is localized in the cytoplasm, we conclude that the processing of the bacteriocin LcnA to its mature form takes place at the cytosolic side of the cytoplasmic membrane.
Highlights
Many non-lantibiotic bacteriocins of lactic acid bacteria are produced as precursors with N-terminal leader peptides different from those present in preproteins exported by the general sec-dependent secretion pathway
The N-terminal parts of the ATP-binding cassette (ABC) transporter proteins involved in the secretion of the bacteriocins pediocin PA-1 and lactococcin G have been shown to process the precursors of the respective bacteriocins into their mature forms
An even number of transmembrane segments (TMSs) should span the cytoplasmic membrane. Both the PhoA* and LacZ studies indicate that LcnC contains four TMSs in LcnC, but they differ in the exact positioning of these domains
Summary
(Received for publication, October 1, 1998, and in revised form, November 30, 1998). From the Department of Genetics, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands. LcnD is an accessory protein with similarities to other proteins known or believed to be involved in the secretion of various (poly)peptides They always operate in conjunction with a protein from the family of ATP-binding cassette (ABC) transporters. CvaA, a member of the membrane fusion protein family, involved in the secretion of colicin V, has been shown to interact with both a cytoplasmic membrane protein (the ABC transporter) and a protein present in the outer membrane [12] The function of these accessory proteins in Gram-positive bacteria is unknown. The N-terminal parts of the ABC transporter proteins involved in the secretion of the bacteriocins pediocin PA-1 and lactococcin G have been shown to process the precursors of the respective bacteriocins into their mature forms. The results indicate that four membrane helices of LcnC span the cytoplasmic membrane and that both the N- and C termini are cytoplasmic
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