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Background   Pectobacterium carotovorum  subsp.  carotovorum  belongs to the  Enterobacteriaceae  family, which causes soft-rot disease in numerous plants worldwide resulting in significant economic losses. Results from our previous studies showed that the strain H-rif-8-6 produces low-molecular-weight bacteriocin (LMWB) Carocin S1. Interestingly, TH22–10, the  caroS1K:Tn5  insertional mutant in H-rif-8-6, loses Carocin S1 producing ability, but still produces other LMWBs which the indicator strain SP33 can detect. The SP33 is one of the many strains that are sensitive toward the cytotoxic effects of Carocin S3K, but not Carocin S1. The result revealed that H-rif-8-6 is a multiple-bacteriocin producing strain.    Results  In this study, a 4.1-kb DNA fragment was isolated from the chromosomal DNA of Pcc strain, H-rif-8-6, by a DNA probe using the  caroS1K  gene as the template. DNA sequencing and analysis by GenBank revealed two complete open reading frames (ORFs), designated ORF1 and ORF2, which were identified within the sequence fragment. ORF1 and ORF2, similar to the identified  carocin S2  genes, encode the killer (Carocin S3K) and the immunity (Carocin S3I) proteins, respectively, which were homologous to the  colicin E3  gene. Carocin S3K and Carocin S3I were expressed, isolated, and purified in  Escherichia coli  BL21 after subcloning of the expression plasmid pGS3KI or pGSK3I. SDS-PAGE analysis showed that the relative masses of Carocin S3K and Carocin S3I were 95.6 kDa and 10.2 kDa, respectively. The results reveal that Carocin S3K has higher antimicrobial and specific antimicrobial activities for Pcc along with a nuclease activity than Carocin S3I. However, Carocin S3I inhibits the activity of Carocin S3K. Interestingly, a high concentration of Carocin S3I protein is also a DNA nuclease, and Carocin S3K also inhibits its activity.    Conclusion  This study showed that another type of bacteriocin was found in  Pectobacterium carotovorum.  This new type of bacteriocin, Carocin S3, has the killer protein, Carocin S3K, and the immunity protein, Carocin S3I.

bmc microbiology

Identification and characterization of the bacteriocin Carocin S3 from the multiple bacteriocin producing strain of Pectobacterium carotovorum subsp. carotovorum