PURIFICATION AND CLONING OF SAKACIN-674, A BACTERIOCIN FROM LACTOBACILLUS-SAKE LB674

Sakacin 674, a bacteriocin produced by Lactobacillus sake Lb764 and wh ich inhibits the growth of Listeria monocytogenes, was purified to hom ogeneity by ammonium sulphate precipitation and sequential ion exchang e, hydrophobic interaction and reversed phase chromatography. The comp lete amino acid sequence of sakacin 674 was determined by Edman degrad ation. The bacteriocin consisted of 43 amino acid residues and had a c alculated molecular mass of 4436.6 Da, which is in good agreement with the molecular mass of 4437.2 as determined by mass spectrometry. The structural gene encoding sakacin 674 (sakR) was located on the chromos ome. This gene was cloned and sequenced. It encoded a primary translat ion product of 61 amino acid residues which was cleaved between amino acids 18 and 19 to yield the active sakacin 674. Sakacin 674 resembled other known bacteriocins and was very similar to sakacin P.