Abstract
BackgroundPediocin-like bacteriocins, ribosomally-synthesized antimicrobial peptides, are generally coexpressed with cognate immunity proteins in order to protect the bacteriocin-producer from its own bacteriocin. As a step for understanding the mode of action of immunity proteins, we determined the crystal structure of PedB, a pediocin-like immunity protein conferring immunity to pediocin PP-1.ResultsThe 1.6 Å crystal structure of PedB reveals that PedB consists of an antiparallel four-helix bundle with a flexible C-terminal end. PedB shows structural similarity to an immunity protein against enterocin A (EntA-im) but some disparity to an immunity protein against carnobacteriocin B2 (ImB2) in both the C-terminal conformation and the local structure constructed by α3, α4, and their connecting loop. Structure-inspired mutational studies reveal that deletion of the last seven residues of the C-terminus of PedB almost abolished its immunity activity.ConclusionThe fact that PedB, EntA-im, and ImB2 share a four-helix bundle structure strongly suggests the structural conservation of this motif in the pediocin-like immunity proteins. The significant difference in the core structure and the C-terminal conformation provides a structural basis for the classification of pediocin-like immunity proteins. Our mutational study using C-terminal-shortened PedBs and the investigation of primary sequence of the C-terminal region, propose that several polar or charged residues in the extreme C-terminus of PedB which is crucial for the immunity are involved in the specific recognition of pediocin PP-1.
Highlights
Pediocin-like bacteriocins, ribosomally-synthesized antimicrobial peptides, are generally coexpressed with cognate immunity proteins in order to protect the bacteriocinproducer from its own bacteriocin
NMR studies revealed that type IIa bacteriocins consists of two domains, a cationic N-terminal domain and a hydrophobic C-terminal domain [10,11,12]: the N-terminal domain interacts with the anionic cell surface of Grampositive bacteria [13,14], while the C-terminal domain participates in the membrane permeabilization [7,15,16]
To investigate whether PedB confers immunity to pediocin PP-1, we introduced the pedB gene into the bacteriocin-sensitive Lactobacillus sakei NCFB 2714 strain [24] and tested the susceptibility against ammonium sulphate-precipitated fermentate of P. pentosaceus containing pediocin PP-1 (See Methods)
Summary
Pediocin-like bacteriocins, ribosomally-synthesized antimicrobial peptides, are generally coexpressed with cognate immunity proteins in order to protect the bacteriocinproducer from its own bacteriocin. Bacteriocins are ribosomally-synthesized antimicrobial peptides produced by bacteria. Most bacteriocins are generally synthesized as prepeptides and can be classified into several classes depending on their post-translational processing and the mode of action [1,2,3]. Type IIa bacteriocins are matured by simple cleavage of a leader peptide and characterized by a conserved YGNGVXC motif in the N-terminus [5,6,7]. NMR studies revealed that type IIa bacteriocins consists of two domains, a cationic N-terminal domain and a hydrophobic C-terminal domain [10,11,12]: the N-terminal domain interacts with the anionic cell surface of Grampositive bacteria [13,14], while the C-terminal domain participates in the membrane permeabilization [7,15,16]
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