Abstract
BackgroundMost isolates of Pectobacterium carotovorum subsp. carotovorum (Pcc) produce bacteriocins. In this study, we have determined that Pcc strain F-rif-18 has a chromosomal gene encoding the low-molecular-weight bacteriocin, Carocin S2, and that this bacteriocin inhibits the growth of a closely related strain. Carocin S2 is inducible by ultraviolet radiation but not by mutagenic agents such as mitomycin C.ResultsA carocin S2-defective mutant, TF1-2, was obtained by Tn5 insertional mutagenesis using F-rif-18. A 5706-bp DNA fragment was detected by Southern blotting, selected from a genomic DNA library, and cloned to the vector, pMS2KI. Two adjacent complete open reading frames within pMS2KI were sequenced, characterized, and identified as caroS2K and caroS2I, which respectively encode the killing protein and immunity protein. Notably, carocin S2 could be expressed not only in the mutant TF1-2 but also in Escherichia coli DH5α after entry of the plasmid pMS2KI. Furthermore, the C-terminal domain of CaroS2K was homologous to the nuclease domains of colicin D and klebicin D. Moreover, SDS-PAGE analysis showed that the relative mass of CaroS2K was 85 kDa and that of CaroS2I was 10 kDa.ConclusionThis study shown that another nuclease type of bacteriocin was found in Pectobacterium carotovorum. This new type of bacteriocin, Carocin S2, has the ribonuclease activity of CaroS2K and the immunity protein activity of CaroS2I.
Highlights
Most isolates of Pectobacterium carotovorum subsp. carotovorum (Pcc) produce bacteriocins
Mutant colonies were identified by smaller inhibition zones. This evidence of mutation suggested that transposon Tn5 had been inserted into LMW bacteriocin-related genes
Isolates of Pcc were distinguished from Escherichia coli by their ability to grow on Modified Drigalski’s agar of the immunity protein, CaroS2I, is usually to stop the damage caused by CaroS2K in the cytoplasm
Summary
Most isolates of Pectobacterium carotovorum subsp. carotovorum (Pcc) produce bacteriocins. Pcc produces many extracellular pectic enzymes (pectate lyase, pectin lyase, exopolygalacturnoate lyase) and hydrolytic enzymes causing soft-rot disease, tissue maceration, and cell wall collapse [2,3]. The only current strategy against soft-rot disease involves chemical agents that unavoidably contaminate the environment [4]. Kikumoto et al have demonstrated that mixed bacteriocin-producing avirulent strains of Pcc show high efficacy against soft-rot disease of Chinese cabbage [5]. Bacteriocins are bactericidal, extracellular toxins, produced by both Gram-positive and Gram-negative bacteria [6,7]. These proteinaceous molecules kill closely related bacteria. Bacteriocins (especially nuclease bacteriocins) have a high amino acid sequence homology
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