Characteristics and genetic determinant of a hydrophobic peptide bacteriocin, carnobacteriocin A, produced by Carnobacterium piscicola LV17A.
Carnobacteriocin A is a hydrophobic nonlantibiotic bacteriocin that is detected early in the growth cycle of Carnobacterium piscicola LV17A and encoded by a 49 MDa plasmid. The bacteriocin was purified using hydrophobic interaction and gel filtration chromatography, and reversed-phase HPLC. Three different active peaks (A1, A2 and A3) were detected, but the purified samples had identical N-terminal amino acid sequences for the first 15 amino acids as determined by Edman degradation analysis. Only a 2.4 kb fragment of the EcoRI digest of the plasmid pCP49 hybridized with a 23-mer oligonucleotide probe derived from amino acids 5 to 13 of the amino acid sequence. The structural gene for carnobacteriocin A is located 600 base pairs into the 2.4 kb EcoRI fragment, but no other genetic information was detected on this unit. The structural gene includes an 18 amino acid N-terminal extension of the bacteriocin, ending with Gly-Gly residues in the -2, -1 positions with respect to the cleavage site. The bacteriocin consists of 53 amino acids that differ markedly from the majority of hydrophobic peptide bacteriocins characterized to date. Based on the amino acid sequence derived from the nucleotide sequence a molecular mass of 5052.85 Da was calculated. Mass spectrometric analysis showed that the molecular mass of the major component (A3) was 2 Da lower, thereby indicating the presence of a disulphide bridge between Cys 22 and Cys 51. Carnobacteriocin A2 has a similar structure except that Met 52 is oxidized to a sulphoxide, whereas A1 appears to be a mixture of peptides derived proteolytically from A3 or A2.
Department of Chemistry, University of Alberta, Edmonton, Canada.
R W
RW
Worobo
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T
Henkel
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M
Sailer
K L
KL
Roy
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JC
Vederas
M E
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Stiles
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics*; D001430:Bacteriocins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics; D004355:Drug Stability; D005516:Food Microbiology; D005798:Genes, Bacterial; D006093:Gram-Positive Asporogenous Rods / Q000235:genetics* / Q000254:growth & development / Q000302:isolation & purification; D008460:Meat / Q000382:microbiology; D008969:Molecular Sequence Data; D008970:Molecular Weight
D001426:Bacterial Proteins; D001430:Bacteriocins; D004269:DNA, Bacterial; C087892:cbnBA protein, Carnobacterium piscicola