Cloning of hybrid bacteriocins with different hinge regions

NAVARRO, S.; ACUÑA, L; GALVAN, AE; BELLOMIO, A; CHALON M.C.

Mar del Plata

Congreso; Reunión Anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2015

SAIB

The ribosomally synthesized antimicrobial peptides produced by bacteria are called bacteriocins. Generally the onesproduced by Gram (-) are active against other Gram (-), in the same way, the ones secreted by Gram (+) bacteria areactive against other Gram (+) phylogenetically related to the producing strain. The aim of this work was to producehybrid bacteriocins active against both kinds of bacteria. To achieve it, we conducted transcriptional fusions betweenthe structural genes of different bacteriocins produced by lactic acid bacteria, and the gen of microcin V (MccV), abacteriocin produced by Escherichia coli. Genes were fused by asymmetric PCR reactions, intercalating differentcoding sequences for a region called "hinge", in order to obtain broad spectrum recombinant chimerical bacteriocins.The fused genes were cloned into the expression vector pET 28 to produce the fusion proteins. All constructs wereexpressed in E. coli BL21 (DE3) after IPTG induction. The antimicrobial activity of different cell extracts andpartially purified preparations of hybrid bacteriocins were tested against Gram (-) and Gram (+) reporter strains