Mechanistic and structural basis of bioengineered bovine Cathelicidin-5 with optimized therapeutic activity

Bikash R Sahoo,Toshimichi Fujiwara,Yuxi Lin,Jyotheeswara R Edula,Kenta Maruyama,Young-Ho Lee,Takahiro Tougan,Toshihiro Horii

doi:10.1038/srep44781

Abstract

Peptide-drug discovery using host-defense peptides becomes promising against antibiotic-resistant pathogens and cancer cells. Here, we customized the therapeutic activity of bovine cathelicidin-5 targeting to bacteria, protozoa, and tumor cells. The membrane dependent conformational adaptability and plasticity of cathelicidin-5 is revealed by biophysical analysis and atomistic simulations over 200 μs in thymocytes, leukemia, and E. coli cell-membranes. Our understanding of energy-dependent cathelicidin-5 intrusion in heterogeneous membranes aided in designing novel loss/gain-of-function analogues. In vitro findings identified leucine-zipper to phenylalanine substitution in cathelicidin-5 (1–18) significantly enhance the antimicrobial and anticancer activity with trivial hemolytic activity. Targeted mutants of cathelicidin-5 at kink region and N-terminal truncation revealed loss-of-function. We ensured the existence of a bimodal mechanism of peptide action (membranolytic and non-membranolytic) in vitro. The melanoma mouse model in vivo study further supports the in vitro findings. This is the first structural report on cathelicidin-5 and our findings revealed potent therapeutic application of designed cathelicidin-5 analogues.

Highlights

Antibiotic treatment of invading pathogens is one of the promising solution to manage an array of invasive microbial infections
The cytotoxic activity of BMAP-28 is greater than its homologues proteins BMAP-27 and LL-37 which suggests its potential therapeutic advantages and disadvantages[14,16,17]
We explored the mechanism of BMAP-28 binding to heterogeneous membranes that mimics bacteria, leukemia, and thymocyte cells at atomistic resolution

Summary

Introduction

Antibiotic treatment of invading pathogens is one of the promising solution to manage an array of invasive microbial infections. Major of the AMPs target the pathogenic cells by membranolytic mechanism, while few show non-membranolytic and intracellular activity of action[9,10,11]. A profound study is needed to understand their mechanism of action which could help in designing potent analogues to act on multiple diseases To this end, another major factor i.e. the plasma membrane heterogeneity driving BMAP-28 action need to be explored. The selective toxicity of BMAP-28 can be considered as an ideal target to explore the mechanism of action through molecular optimization To address these limitations, we explored the mechanism of BMAP-28 binding to heterogeneous membranes that mimics bacteria, leukemia, and thymocyte cells at atomistic resolution. The therapeutic activity of BMAP-28 and its optimized derivatives were further investigated in vitro and in vivo to ensure the cell and/or organism specific cytotoxic activity

Methods

Results

Conclusion