Expression of Caseicin from Lacticaseibacillus casei and Lacticaseibacillus zeae Provides Insight into Antilisterial Class IIa Bacteriocins

• Published in: Probiotics and antimicrobial proteins 2024-08
• Main Authors: Salini, Francesco, Vermeulen, Ross, du Preez van Staden, Anton, Comi, Giuseppe, Iacumin, Lucilla, Dicks, Leon M T
• Format: Article
• Language: English
• ISSN: 1867-1306; 1867-1314; 1867-1314
• DOI: 10.1007/s12602-024-10341-0

In this study, an in silico screening approach was employed to mine potential bacteriocin clusters in genome-sequenced isolates of Lacticaseibacillus zeae UD 2202 and Lacticaseibacillus casei UD 1001. Two putative undescribed bacteriocin gene clusters (Cas1 and Cas2) closely related to genes encoding class IIa bacteriocins were identified. No bacteriocin activity was recorded when cell-free supernatants of strains UD 2202 and UD 1001 were tested against Listeria monocytogenes. Genes encoding caseicin A1 (casA1) and caseicin A2 (casA2) were heterologously expressed in Escherichia coli BL21 (DE3) using the nisin leader peptide cloned in-frame to the C-terminal of the green fluorescent gene (mgfp5). Nisin protease (NisP) was used to cleave caseicin A1 (casA1) and caseicin A2 (casA2) from GFP-Nisin leader fusion proteins. Both heterologously expressed peptides (casA1 and casA2) inhibited the growth of L. monocytogenes, suggesting that casA1 and casA2 are either silent in the wild-type strains or are not secreted in an active form. The minimum inhibitory concentration (MIC) of casA1 and casA2, determined using HPLC-purified peptides, ranged from