TY - JOUR
T1 - The circular bacteriocin enterocin NKR-5-3B has an improved stability profile over nisin
AU - Wang, Conan K.
AU - Huang, Yen Hua
AU - Shabbir, Fatima
AU - Pham, Huong T.
AU - Lawrence, Nicole
AU - Benfield, Aurélie H.
AU - van der Donk, Wilfred
AU - Henriques, Sónia T.
AU - Turner, Mark S.
AU - Craik, David J.
N1 - This work was supported by access to the facilities of the Australian Research Council (ARC) Centre of Excellence for Innovations in Peptide and Protein Science ( CE200100012 ). CKW is an ARC Future Fellow ( FT220100583 ). DJC is a National Health and Medical Research Council (NHMRC) Leadership Fellow (grant number 2009564 ). NL is supported by NHMRC grant 1183927 . STH and AB are supported by ARC grant CE200100012 .
PY - 2023/9
Y1 - 2023/9
N2 - Bacteriocins are a large family of bacterial peptides that have antimicrobial activity and potential applications as clinical antibiotics or food preservatives. Circular bacteriocins are a unique class of these biomolecules distinguished by a seamless circular topology, and are widely assumed to be ultra-stable based on this constraining structural feature. However, without quantitative studies of their susceptibility to defined thermal, chemical, and enzymatic conditions, their stability characteristics remain poorly understood, limiting their translational development. Here, we produced the circular bacteriocin enterocin NKR-5-3B (Ent53B) in mg/L quantities using a heterologous Lactococcus expression system, and characterized its thermal stability by NMR, chemical stability by circular dichroism and analytical HPLC, and enzymatic stability by analytical HPLC. We demonstrate that Ent53B is ultra-stable, resistant to temperatures approaching boiling, acidic (pH 2.6) and alkaline (pH 9.0) conditions, the chaotropic agent 6 M urea, and following incubation with a range of proteases (i.e., trypsin, chymotrypsin, pepsin, and papain), conditions under which most peptides and proteins degrade. Ent53B is stable across a broader range of pH conditions and proteases than nisin, the most widely used bacteriocin in food manufacturing. Antimicrobial assays showed that differences in stability correlated with differences in bactericidal activity. Overall, this study provides quantitative support for circular bacteriocins being an ultra-stable class of peptide molecules, suggesting easier handling and distribution options available to them in practical applications as antimicrobial agents.
AB - Bacteriocins are a large family of bacterial peptides that have antimicrobial activity and potential applications as clinical antibiotics or food preservatives. Circular bacteriocins are a unique class of these biomolecules distinguished by a seamless circular topology, and are widely assumed to be ultra-stable based on this constraining structural feature. However, without quantitative studies of their susceptibility to defined thermal, chemical, and enzymatic conditions, their stability characteristics remain poorly understood, limiting their translational development. Here, we produced the circular bacteriocin enterocin NKR-5-3B (Ent53B) in mg/L quantities using a heterologous Lactococcus expression system, and characterized its thermal stability by NMR, chemical stability by circular dichroism and analytical HPLC, and enzymatic stability by analytical HPLC. We demonstrate that Ent53B is ultra-stable, resistant to temperatures approaching boiling, acidic (pH 2.6) and alkaline (pH 9.0) conditions, the chaotropic agent 6 M urea, and following incubation with a range of proteases (i.e., trypsin, chymotrypsin, pepsin, and papain), conditions under which most peptides and proteins degrade. Ent53B is stable across a broader range of pH conditions and proteases than nisin, the most widely used bacteriocin in food manufacturing. Antimicrobial assays showed that differences in stability correlated with differences in bactericidal activity. Overall, this study provides quantitative support for circular bacteriocins being an ultra-stable class of peptide molecules, suggesting easier handling and distribution options available to them in practical applications as antimicrobial agents.
KW - Antimicrobial peptide
KW - Stability
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U2 - 10.1016/j.peptides.2023.171049
DO - 10.1016/j.peptides.2023.171049
M3 - Article
C2 - 37390898
AN - SCOPUS:85164290805
SN - 0196-9781
VL - 167
JO - Peptides
JF - Peptides
M1 - 171049
ER -