Gene encoded antimicrobial peptides, a template for the design of novel anti-mycobacterial drugs

Abstract

Nisin A is the most widely characterized lantibiotic investigated to date. It represents one of the many antimicrobial peptides which have been the focus of much interest as potential therapeutic agents. This has resulted in the search for novel lantibiotics and more commonly, the engineering of novel variants from existing peptides with a view to increasing their activity, stability and solubility.The aim of this study was to compare the activities of nisin A and novel bioengineered hinge derivatives, nisin S, nisin T and nisin V. The microtitre alamar blue assay (MABA) was employed to identify the enhanced activity of these novel variants against M. tuberculosis (H37Ra), M. kansasii (CIT11/06), M. avium subsp. hominissuis (CIT05/03) and M. avium subsp. paratuberculosis (MAP) (ATCC 19698). All variants displayed greater anti-mycobacterial activity than nisin A. Nisin S was the most potent variant against M. tuberculosis, M. kansasii and M. avium subsp. hominissuis, retarding growth by a maximum of 29% when compared with nisin A. Sub-species variations of inhibition were also observed with nisin S reducing growth of Mycobacterium avium subsp. hominissuis by 28% and Mycobacterium avium subsp. paratuberculosis by 19% and nisin T contrastingly reducing growth of MAP by 27% and MAC by 16%.Nisin S, nisin T and nisin V are potent novel anti-mycobacterial compounds, which have the capacity to be further modified, potentially generating compounds with additional beneficial characteristics. This is the first report to demonstrate an enhancement of efficacy by any bioengineered bacteriocin against mycobacteria.

Keywords: alamar blue; bacteriocin; lantibiotic; mycobacteria; nisin variants; peptide engineering.

Figure 1

Structure of nisin variants Nisin A, Nisin V, Nisin S and Nisin T. Dark circles indicate amino acid differences between Nisin A and its derivatives.

Figure 2

Agar diffusion assay of Nisin A and its derivatives Nisin V, Nisin T and Nisin S, overlaid with M. smegmatis.

Figure 3

Activity of nisin V (M21V), nisin T (K22T), nisin S (K22S) and wild-type nisin A against a cohort of pathogenic mycobacteria. Efficacy was evaluated using alamar blue colorimetric indicator dye. The dye reduction value of nisin A was taken as 100% relative growth, facilitating direct comparison of nisin A to the activity of nisin V, nisin T and nisin S.