A SIGNAL PEPTIDE SECRETION-DEPENDENT BACTERIOCIN FROM CARNOBACTERIUM DIVERGENS

Divergicin A is a strongly hydrophobic, narrow-spectrum, nonlantibioti c bacteriocin produced by Carnobacterium divergens LV13. This strain o f C. divergens contains a 3.4-kb plasmid that mediates production of, and immunity to, the bacteriocin, N-terminal amino acid sequencing of the purified divergicin A was used to locate the structural gene (dvnA ). The structural gene encodes a prepeptide of 75 amino acids consisti ng of a 29-amino-acid N-terminal extension and a mature peptide of 46 amino acids, Directly downstream of dvnA there is a second open readin g frame that encodes the immunity protein for divergicin A. Divergicin A has a calculated molecular mass of 4,223.89 Da. The molecular mass determined by mass spectrometry is 4,223.9 Da, indicating that there i s no posttranslational modification of the peptide, The N-terminal ext ension of divergicin A has an Ala-Ser-Ala (positions -3 to -1) cleavag e site and acts as a signal peptide that accesses the general export s ystem of the cell (such as the sec pathway in Escherichia coli), This is the first bacteriocin of lactic acid bacteria to be reported that d oes not have dedicated maturation and secretion genes, Production of d ivergicin A was observed in heterologous hosts containing only the two genes associated with divergicin A production and immunity, Fusing al kaline phosphatase behind the signal peptide for divergicin resulted i n the secretion of this enzyme in the periplasmic space and supernatan t of E. coli.