Molecular and Genetic Characterization of Propionicin F, a Bacteriocin fromPropionibacterium freudenreichii

Abstract

This work describes the purification and characterization of propionicin F, the first bacteriocin isolated from Propionibacterium freudenreichii. The bacteriocin has a bactericidal activity and is only active against strains of P. freudenreichii. Propionicin F appears to be formed through a processing pathway new to bacteriocins. The mass of the purified bacteriocin was determined by mass spectrometry, and the N-terminal amino acid sequence was determined by Edman degradation. Sequencing of pcfA, the bacteriocin structural gene, revealed that propionicin F corresponds to a 43-amino-acid peptide in the central part of a 255-amino-acid open reading frame, suggesting that mature propionicin F is excised from the probacteriocin by N-and C-terminal proteolytic modifications. DNA sequencing and Northern blot hybridizations revealed that pcfA is cotranscribed with genes encoding a putative proline peptidase and a protein from the radical S-adenosylmethionine family. A gene encoding an ABC transporter was also identified in close proximity to the bacteriocin structural gene. The potential role of these genes in propionicin F maturation and secretion is discussed.Propionic acid bacteria (PAB) and lactic acid bacteria (LAB) have been utilized for flavor development and preservation of food products for centuries. The preservative capacity of PAB and LAB has previously been attributed to the lowering of pH by production of propionic acid, acetic acid, and lactic acid, which are the main fermentative end products produced by these organisms. However, it has become evident that PAB and LAB also produce antimicrobial substances in addition to organic acids (25). Among these are the bacteriocins, which are gene-encoded, ribosomally synthesized peptides that usually display antimicrobial activity against species closely related to the producer organism (43).A vast number of LAB bacteriocins have been purified; some of these, including nisin, pediocin PA-1, and sakacin P, have been thoroughly characterized at the genetic and biochemical level (5,6,16,21,24,26,27,29,33,40,44,45). In general, LAB bacteriocins are small, cationic peptides, with a propensity to adopt amphiphilic helical structures (36). There are, however, exceptions from this rule, including helveticin J and enterolysin A, which both are large proteins with antimicrobial activity (23, 37). The LAB bacteriocins are usually synthesized as prepeptides, containing an N-terminal secretion signal peptide of the sec or the double-glycine type. The leader sequence is cleaved off concomitant with secretion either by the general secretory machinery or, in the case of doubleglycine leaders, by dedicated ABC transporters (15,47).In contrast to the numerous bacteriocins characterized from LAB, only three PAB bacteriocins have been identified and characterized at the amino acid and genetic level. Propionicin T1 isolated from Propionibacterium thoenii is a 65-amino-acid bacteriocin, synthesized with a 31-residue signal peptide (13). The 204-amino-acid bacteriocin SM1 from Propion...

Thuricin CD, a posttranslationally modified bacteriocin with a narrow spectrum of activity against Clostridium difficile

“…TrnC exhibited sequence similarity to conserved domains of the Fe-S oxidoreductase arylsulfatase regulatory proteins (AslB), and TrnD had sequences similar to the MoaA family of molybdenum cofactor enzymes. Radical SAM enzymes rarely occur in bacteriocin gene clusters, but they are included in those of subtilosin A and propiocin F (26,27). Sequence alignments indicated that AlbA of subtilosin A exhibits only 19% and 17% identity to TrnC and TrnD, respectively, although AlbA exhibits some features of each individual thuricin CD protein.…”

Dairy Propionibacteria: Less Conventional Probiotics to Improve the Human and Animal Health

“…Among them it could be mentioned: Propionicin PLG-1 and GBZ-1 produced by P. thoenii 127 [81]; Jenseniin G isolated from P. thoenii P126 [82]; Propionicins SM1 and SM2 produced by P. jensenii DF1 [83]; Propionicin T1 synthesized by P. thoenii 419 and LMG2792 [84]; Thoenicin 447 isolated from P. thoenii 447 [85]; Acnecin produced by a strain of P. acnes [86] and several other propionicins [87][88][89].…”

Heterologous Production of Antimicrobial Peptides in Propionibacterium freudenreichii

“…Electroporation of competent P. freudenreichii subsp. shermanii IFO12426 cells produced 10 4 and Ͼ10 7 transformants/g DNA with vector prepared from E. coli and P. freudenreichii, respectively. With DNA from E. coli, this is an improvement in transformation efficiency by 3 orders magnitude compared to the transformation of P. freudenreichii ATCC 6207.…”

“…PAB also have a potential use as protective cultures for inhibition of pathogens and food spoilage organisms (30,31). The antimicrobial capacity of PAB is only partly due to the production of organic acids, and it has become evident that PAB also produce other biologically active substances such as bacteriocins (4,7,9,13,26). Recently, Faye et al (9) characterized the propionicin T1 bacteriocin from Propionibacterium thoenii.…”