Analysis of the sakacin P gene cluster from Lactobacillus sake Lb674 and its expression in sakacin-negative Lb. sake strains.
Sakacin P is a small, heat-stable, ribosomally synthesized peptide produced by certain strains of Lactobacillus sake. It inhibits the growth of several Gram-positive bacteria, including Listeria monocytogenes. A 7.6 kb chromosomal DNA fragment from Lb. sake Lb674 encompassing all genes responsible for sakacin P production and immunity was sequenced and introduced into Lb. sake strains Lb790 and Lb706X which are bacteriocin-negative and sensitive to sakacin P. The transformants produced sakacin P in comparable amounts to the parental strain, Lb674. The sakacin P gene cluster comprised six consecutive genes: sppK, sppR, sppA, spiA, sppT and sppE, all transcribed in the same direction. The deduced proteins SppK and SppR resembled the histidine kinase and response regulator proteins of bacterial two-component signal transducing systems of the AgrB/AgrA-type. The genes sppA and spiA encoded the sakacin P preprotein and the putative immunity protein, respectively. The predicted proteins SppT and SppE showed strong similarities to the proposed transport proteins of several other bacteriocins and to proteins implicated in the signal-sequence-independent export of Escherichia coli haemolysin A. Deletion and frameshift mutation analyses showed that sppK, sppT and sppE were essential for sakacin P production in Lb706X. The putative SpiA peptide was shown to be involved in immunity to sakacin P. Analogues of sppR and spiA were found on the chromosomes of Lb. sake Lb706X and Lb790, indicating the presence of an incomplete spp gene cluster in these strains.
Hühne
Kathrin
K
Institute for Microbiology and Toxicology, Federal Center of Meat Research, E.-C.-Baumann-Straβe 20, D-95326 Kulmbach, Germany.
Axelsson
Lars
L
MATFORSK, Norwegian Food Research Institute, Osloveien 1, N-1430 Ås, Norway.
Holck
Askild
A
MATFORSK, Norwegian Food Research Institute, Osloveien 1, N-1430 Ås, Norway.
Kröckel
Lothar
L
Institute for Microbiology and Toxicology, Federal Center of Meat Research, E.-C.-Baumann-Straβe 20, D-95326 Kulmbach, Germany.
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001430:Bacteriocins / Q000096:biosynthesis / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D005798:Genes, Bacterial; D000071677:Histidine Kinase; D007778:Lactobacillus / Q000235:genetics*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011494:Protein Kinases / Q000235:genetics; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D015398:Signal Transduction; D013045:Species Specificity; D014169:Transformation, Bacterial
D001426:Bacterial Proteins; D001430:Bacteriocins; D004269:DNA, Bacterial; C087217:sakacin P protein, Lactobacillus; D011494:Protein Kinases; D000071677:Histidine Kinase