Characterization and cloning of the genes encoding enterocin 1071A and enterocin 1071B, two antimicrobial peptides produced by Enterococcus faecalis BFE 1071.
The pH-neutral cell supernatant of Enterococcus faecalis BFE 1071, isolated from the feces of minipigs in Göttingen, inhibited the growth of Enterococcus spp. and a few other gram-positive bacteria. Ammonium sulfate precipitation and cation-exchange chromatography of the cell supernatant, followed by mass spectrometry analysis, yielded two bacteriocin-like peptides of similar molecular mass: enterocin 1071A (4.285 kDa) and enterocin 1071B (3.899 kDa). Both peptides are always isolated together. The peptides are heat resistant (100 degrees C, 60 min; 50% of activity remained after 15 min at 121 degrees C), remain active after 30 min of incubation at pH 3 to 12, and are sensitive to treatment with proteolytic enzymes. Curing experiments indicated that the genes encoding enterocins 1071A and 1071B are located on a 50-kbp plasmid (pEF1071). Conjugation of plasmid pEF1071 to E. faecalis strains FA2-2 and OGX1 resulted in the expression of two active peptides with sizes identical to those of enterocins 1071A and 1071B. Sequencing of a DNA insert of 9 to 10 kbp revealed two open reading frames, ent1071A and ent1071B, which coded for 39- and 34-amino-acid peptides, respectively. The deduced amino acid sequence of the mature Ent1071A and Ent1071B peptides showed 64 and 61% homology with the alpha and beta peptides of lactococcin G, respectively. This is the first report of two new antimicrobial peptides representative of a fourth type of E. faecalis bacteriocin.
Balla
E
E
Department of Microbiology, University of Stellenbosch, Stellenbosch 7600, South Africa.
Dicks
L M
LM
Du Toit
M
M
Van Der Merwe
M J
MJ
Holzapfel
W H
WH
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D001430:Bacteriocins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology*; D001483:Base Sequence; D003001:Cloning, Molecular; D003227:Conjugation, Genetic; D013293:Enterococcus faecalis / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D005798:Genes, Bacterial; D006094:Gram-Positive Bacteria / Q000187:drug effects*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000378:metabolism; D010957:Plasmids; D017422:Sequence Analysis, DNA
D001426:Bacterial Proteins; D001430:Bacteriocins; D010447:Peptide Hydrolases