Novel antimicrobial peptide derived from the fusion of bacteriocins from Gram (+) and Gram (-) bacteria

ACUÑA L; NIKLISON CHIROU MV; SESMA F; MORERO RD; BELLOMIO A

Mar del Plata, Buenos Aires. Argentina

Congreso; SAIB XLIII Reunión Anual; 2007

SAIB

Enterocin CRL35 is a class IIa bacteriocin produced by Enterococcus mundtii, a Gram (+) bacterium, while colicin V is a microcin produced by the Gram (-) E. coli. Both antimicrobial peptides are active against their closely-related microorganisms. The aim of this work was to fuse the structural gene of enterocin CRL35 (munA) and colicinV(cvaC) in order to obtain recombinant fusion proteins, namely MunA-CvaC and CvaC-MunA with antimicrobial activity of wider antimicrobial spectrum. Since these peptides do not have postranslational modifications, the construction of a hybrid antimicrobial peptide was possible. The structural genes of enterocin CRL35 and colicin V were amplified by PCR. Then, they were fused using the megaprimer technique. The amplicons were cloned in the vector pET28b and expressed in E. coli BL21[DE3]. The transforming clones were cultivated in LB media and gene expression was induced with IPTG. Polyacrilamide gel electrophoresis both in native and denaturalizing conditions were performed in order to confirm the correct expression of the hybrids. Antimicrobial activity on the native gel was determined with sensitive strains. A Gram (+) and Gram (-) active fusion peptide Mun35-CvaC was obtained that could be visualized by polyacrilamide gel electrophoresis. These results showed a promising technique for obtaining novel antimicrobial peptides.