RESUMO
OBJECTIVE: Pediocin PA-1, an antimicrobial peptide derived from Pediococcus acidilactici PAC1.0, has a potential application as a food preservative thanks to its strong inhibitory activity against the foodborne pathogen L. monocytogenes. This study aimed to produce Pediocin PA-1 from the yeast P. pastoris and evaluate its characteristics. METHODS: Gene encoding Pediocin PA-1 was integrated into P. pastoris X33 genome to establish the strain X33::ped, which could produce and secrete this peptide into culture medium. The antimicrobial activity of Pediocin PA-1 was examined using agar diffusion assay. The stability of pediocin PA-1 was determined based on its remaining antibacterial activity after exposure to proteases and extreme pH and temperatures. The potential use of this bacteriocin in food preservation was demonstrated using the L. monocytogenes infected pork bologna. The anticancer activity of Pediocin PA-1 was also investigated on some cancer cells using MTT assay. RESULTS: We established the yeast P. pastoris X33::ped capable of producing pediocin PA-1 with antimicrobial activity against L. monocytogenes and some other harmful bacteria. Pediocin PA-1 was stable at 100ËC and resistant against pH 1-12 for 1 h, but susceptible to trypsin, α-chymotrypsin, and proteinase K. This peptide could reduce the number of L. monocytogenes in pork bologna by 3.59 log CFU/g after 7 days of storage at 4ËC. Finally, Pediocin PA-1 (25 µg/ml) inhibited the proliferation of A549 and Hela cancer cells. CONCLUSION: We succeeded in producing active Pediocin PA-1 from P. pastoris and demonstrated its potential use in food preservation and pharmaceutical industry.
Assuntos
Conservação de Alimentos , Listeria monocytogenes , Pediocinas , Pediocinas/farmacologia , Pediocinas/genética , Animais , Conservação de Alimentos/métodos , Listeria monocytogenes/efeitos dos fármacos , Listeria monocytogenes/genética , Humanos , Antibacterianos/farmacologia , Clonagem Molecular , Suínos , Testes de Sensibilidade Microbiana , Bacteriocinas/farmacologia , Bacteriocinas/genética , Bacteriocinas/metabolismo , Pediococcus acidilactici/genética , Pediococcus acidilactici/metabolismo , Expressão Gênica , Saccharomycetales/genética , Saccharomycetales/metabolismo , Saccharomycetales/efeitos dos fármacosRESUMO
In this study, 10 lactic acid bacteria were isolated from Turkish fermented sausage (sucuk) and identified as 5 Lactobacillus plantarum, 1 Pediococcus acidilactici, 1 Weissella hellenica, 1 Lactobacillus pentosus, and 2 Lactobacillus sakei. PCR screening of genes encoding plantaricin A and pediocin showed the presence of plantaricin A gene in 9 and pediocin gene in 3 of strains. All isolates showed antibacterial and antifungal effect on most of the tested microorganisms. gad gene, encoding glutamic acid decarboxylase enzyme, was detected in all isolates except Weisella hellenica KS-24. Eight of isolates were determined as gamma-amino butyric acid (GABA) producer in the presence of 53 mM mono sodium glutamate (MSG) by HPLC and TLC analysis. DPPH scavenging activity was observed for all isolates. Additionally, isolates were able to produce exopolysaccharide in the presence of sucrose. The best exopolysaccharide (EPS) production was achieved with L. plantarum KS-11 and L. pentosus KS-27. As a result, this study characterized some techno-functional properties of LAB isolates from sucuk. It was concluded that the isolates studied have the potential to be used in obtaining functional products in meat industry, as well as strain selection may be effective in providing the desired properties in the product.