An Extracellular Loop of the Mannose Phosphotransferase System Component IIC Is Responsible for Specific Targeting by Class IIa Bacteriocins

Abstract

Class IIa bacteriocins target a phylogenetically defined subgroup of mannose-phosphotransferase systems (man-PTS) on sensitive cells. By the use of man-PTS genes of the sensitive Listeria monocytogenes (mpt) and the nonsensitive Lactococcus lactis (ptn) species to rationally design a series of man-PTS chimeras and site-directed mutations, we identified an extracellular loop of the membrane-located protein MptC that was responsible for specific target recognition by the class IIa bacteriocins.Bacteriocins are small, ribosomally synthesized antimicrobial peptides that normally kill bacteria closely related to the bacteriocin producers, but some also target a wider spectrum of bacteria, including a number of pathogens and food spoilage bacterial species (5, 28). Class IIa (pediocin-like) bacteriocins display a broad antimicrobial spectrum, including important pathogens such as Listeria monocytogenes and Enterococcus faecalis. These peptides consist of 37 to 48 nonmodified amino acids, contain a conserved pediocin-box sequence (Y-G-N-G-V/L) in the N-terminal region, and have defined secondary features in their structure: a cationic ␤ sheet at the conserved N terminus and a helix-containing domain at the less-conserved C terminus (16,30). Class IIa bacteriocins target sensitive cells by using the mannose phosphotransferase system (man-PTS) as a receptor (6,10,17,19,33). This sugar uptake system is the major glucose transporter for many bacteria, particularly Firmicutes and Gammaproteobacteria (39). Each man-PTS complex consists of four structural domains: IIC and IID, represented by two membrane-located proteins, and IIA and IIB, which are normally represented by a single cytoplasmic protein that can form reversible contacts with its membrane-located partners (31).It has previously been shown that coexpression of the IIC and IID genes is needed to confer sensitivity to class IIa bacteriocins as well as to the lactococcal bacteriocin lactococcin A and that the cytoplasmic IIAB partner is not involved in this process (10). However, while lactococcin A (belonging to class IIc) targets only the lactococcal man-PTS, the class IIa bacteriocins target man-PTSs of species of diverse genera (e.g., Listeria, Enterococcus, and Lactobacillus) but somehow not those of the Lactococcus genus (24). This genus specificity has been recognized for almost 2 decades (20,23,26); still, the molecular nature underlying the specificity has remained very enigmatic. In the present report we clarify this issue by demonstrating that these two types of bacteriocins exhibit different binding patterns on their receptors: class IIa bacteriocins specifically interact with a defined region of 40 amino acids in the IIC protein whereas lactococcin A has a more complex interaction involving regions from both IIC and IID. MATERIALS AND METHODSBacterial strain and growth conditions. Lactococcus lactis B488 (10), a man-PTS null mutant derived from L. lactis IL1403, was used as an expression host in this study. Lactococcal clones were grown at 30°C in M17 me...

Interaction with Enzyme IIBMpo(EIIBMpo) and Phosphorylation by Phosphorylated EIIBMpoExert Antagonistic Effects on the Transcriptional Activator ManR of Listeria monocytogenes

“…A knockout mutant carrying an insertion of the Tn917 transposon in this gene was isolated based on its resistance to class IIa bacteriocins (34). Similar resistance, together with the inhibition of glucose transport, has also been reported for listeriae and some other firmicutes as a result of mutations that affect the membrane components of the PTS Man (31,35). In this case, however, the Tn917 insertion occurred not in the man operon but in the region between the presumed Ϫ35 and Ϫ10 promoter sites upstream of lin0142.…”

Genetic characterisation and heterologous expression of leucocin C, a class IIa bacteriocin from Leuconostoc carnosum 4010

“…The membrane-located IIC and IID have both been shown to be necessary for the receptor ). Later, co-expression of IIC and IID conferred strong and similar sensitivity to class IIa bacteriocins with the host which only produces IIC, confirming that IIC is the main determinant of specificity (Kjos et al 2010). An N-terminal extracellular loop comprised of 40 aa in this IIC domain was then identified to be the major determinant of the species-specificity.…”

“…An N-terminal extracellular loop comprised of 40 aa in this IIC domain was then identified to be the major determinant of the species-specificity. The loop thus may function as an interaction site for the recognition of class IIa bacteriocins (Kjos et al 2010). The function of IID is not yet known, but it might indirectly interact with the bacteriocin during the pore formation.…”

A Zn-Dependent Metallopeptidase Is Responsible for Sensitivity to LsbB, a Class II Leaderless Bacteriocin of Lactococcus lactis subsp. lactis BGMN1-5

“…Due to this strong specificity, it is generally believed that most, if not all, bacteriocins recognize specific receptors on target cells. Although numerous bacteriocins from Gram-positive bacteria have been characterized over the years, only a few receptors so far have been reported; these are lipid II recognized by nisin and several other lantibiotics, as well as the class IId bacteriocin lactococcin 972 (3,4,23), man-PTS, targeted by class IIa pediocin-like bacteriocins and the class IId lactococcins A and B (5,24), and maltose-ABC transporter, recognized by the circular bacteriocin garvicin ML (8). The possibility that LsbB employs sugar transporters as receptors was also explored.…”