Here, we report on the biochemical characterization of a new glycosylated bacteriocin (glycocin), ASM1, produced by Lactobacillus plantarum A-1 and analysis of the A-1 bacteriocinogenic genes. ASM1 is 43 amino acids in length with Ser18-O- and Cys43-S-linked N-acetylglucosamine moieties that are essential for its inhibitory activity. Its only close homologue, glycocin F (GccF), has five amino acid substitutions all residing in the flexible C-terminal 'tail' and a lower IC50 (0.9 nm) compared to that of ASM1 (1.5 nm). Asm/gcc genes share the same organization (asmH← →asmABCDE→F), and the asm genes reside on an 11 905-bp plasmid dedicated to ASM1 production. The A-1 genome also harbors a gene encoding a 'rare' bactofencin-type bacteriocin. As more examples of prokaryote S-GlcNAcylation are discovered, the functions of this modification may be understood.
- MeSH
- bakteriální geny genetika MeSH
- bakteriociny chemie genetika metabolismus MeSH
- fylogeneze MeSH
- glykosylace MeSH
- Lactobacillus plantarum chemie genetika MeSH
- novobiocin MeSH
- plazmidy genetika MeSH
- sekvence aminokyselin MeSH
- sekvence nukleotidů MeSH
- sekvenční analýza DNA MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The aim of the present study was to develop an ion-selective electrode method for the continuous determination of the intracellular pH in Lactobacillus plantarum using a small-scale bioreactor. This method employed a salicylate-selective electrode basing on the distribution of salicylic acid across the cytoplasmic membrane. This developed electrode responded to salicylate concentrations above 20 μmol/L with a Nernstian sensitivity. The energized and concentrated cells were added into a thermostated small-scale bioreactor that contained the salicylate anions dissolved in a 100 mmol/L potassium phosphate buffer at different pH values. The changes in salicylate concentration that occurred in the medium containing bacterial suspension were measured as a voltage change. The cells of Lactobacillus plantarum showed maintenance of pH homeostasis at the studied pH ranging from 4.0 to 7.0, and they kept a neutral intracellular pH up to 5.8. The simplicity of the measuring preparation and the relatively low cellular concentration, as well as the advantages of the small-scale bioreactor, lead us to believe that the described method can facilitate the study of the physicochemical factors on the intracellular pH of lactic acid bacteria using a single pH probe in one method.
An agar well diffusion assay (AWDA) was used to isolate a high bacteriocin-producing strain with a broad spectrum of antibacterial activity, strain MXG-68, from Inner Mongolia traditional fermented koumiss. Lactobacillus plantarum MXG-68 was identified by morphological, biochemical, and physiological characteristics and 16S rDNA analysis. The production of antibacterial substance followed a growth-interrelated model, starting at the late lag phase of 4 h and arriving at a maximum value in the middle of the stationary phase at 24 h. Antibacterial activity was abolished or decreased in the presence of pepsin, chymotrypsin, trypsin, proteinase, and papain K. The results showed that antibacterial substances produced by L. plantarum MXG-68 were proteinaceous and could thus be classified as the bacteriocin, named plantaricin MXG-68. The molar mass of plantaricin MXG-68 was estimated to be 6.5 kDa, and the amino acid sequence of its N-terminal was determined to be VYGPAGIFNT. The mode of plantaricin MXG-68 action was determined to be bactericidal. Bacteriocin in cell-free supernatant (CFS) at pH 7 was stable at different temperatures (60 °C, 80 °C, 100 °C, 121 °C for 30 min; 4 °C and - 20 °C for 30 days), as well as at pH 2.0-10.0. Antibacterial activity maintained stable after treatment with organic solvents, surfactants, and detergents but increased in response to EDTA. Response surface methodology (RSM) revealed the optimum conditions of bacteriocin production in L. plantarum MXG-68, and the bacteriocin production in medium optimized by RSM was 26.10% higher than that in the basal MRS medium.
- MeSH
- antibakteriální látky biosyntéza chemie farmakologie MeSH
- bakteriociny biosyntéza chemie farmakologie MeSH
- fylogeneze MeSH
- kinetika MeSH
- kultivační média MeSH
- kumys mikrobiologie MeSH
- Lactobacillus plantarum chemie klasifikace fyziologie MeSH
- molekulová hmotnost MeSH
- potravinářská mikrobiologie MeSH
- RNA ribozomální 16S genetika MeSH
- sekvence aminokyselin MeSH
- stabilita proteinů MeSH
- Staphylococcus účinky léků MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Čína MeSH
