Investigating the bacteriocin library Lactobacillus plantarum A-1 : presented in partial fulfilment of the requirements for the degree of Master of Science in Microbiology at Massey University, Manuwatū Campus, Palmerston North, New Zealand

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Bacteriocins are a highly diverse group of ribosomally synthesised, antimicrobial polypeptides produced by nearly all bacterial and archaeal species. Individual bacteriocins typically exhibit a narrow phylogenetic range of activity, but collectively inhibit a wide range of species through a variety of mechanisms. Glycocins are uncommon bacteriocins with rare, S-linked glycosidic bond;, currently there are only four characterised glycocins. Preliminary characterisation of the bacteriocin ASM1 from Lactobacillus plantarum A-1 was reported by Hata et al. in 2010. ASM1 is structurally similar to GccF, a glycocin produced by L. plantarum KW30. Like GccF, ASM1 has two covalently linked N-acetylglucosamine moieties, one of which is attached through a rare S-glycosidic bond. Due to its structural similarity, it was hypothesised that ASM1 would have similar inhibitory activity to GccF. Experiments showed that the two bacteriocins have almost identical inhibitory activity and both glycocins rely on their GlcNAc moieties to inhibit target cells. The range of species inhibited by ASM1 was shown to be wider than previously thought. The inhibitory activity, however, varied considerably even between strains in a species. The ASM1 gene cluster was established by sequencing and Southern hybridisation to be located on a 11,905 bp plasmid pA1_ASM1. An in silico analysis of the ASM1 gene cluster showed it to have the same operonic organisation similar as the GccF cluster, and overall DNA sequence identity of 75%. A second active bacteriocin gene cluster was detected in the L. plantarum A-1 genome. A synthetic peptide, named ASM2, corresponding to this bacteriocin was partially characterised. ASM2 has 82% amino acid sequence identity to the recently identified bactofencin A produced by L. salivarius DPC6502. A brief in silico analysis of proteins from the A-1 bacteriocin library and their orthologues provided some evolutionary context for the glycocins of Lactobacillus and gave hints as to the evolutionary history of the four characterised glycocins. ASM1 is one of four characterised glycocins and this work has increased the overall knowledge of glycocins. Identification of a novel secondary bacteriocin in L. plantarum A-1 shows the complexity of bacteriocin systems and provides many avenues for future research regarding bacteria that produce multiple bacteriocins.
Bacteriocins, Lactobacillus plantarum