Part:BBa_K1659088

Artilysin Art-E, an antibacterial fusion protein

This part contains the sequence for the proposed novel antimicrobial protein, Art-E.

Introduction

Inspired by the [http://www.ncbi.nlm.nih.gov/pubmed/24752267 work done by Briers et al] on Artilysin Art-175, which our team has catalogued as the BioBrick BBa_K1659000, we created a similar fusion protein using T4 Endolysin via the same design principle of fusing SMAP-29 to an endolysin's N-terminus. T4 Endolysin (more popularly known as T4 Lysozyme) has long been known as an enzyme that can effectively hydrolyze bacterial peptidoglycan, its mechanism of action being the catalytic hydrolysis of the 1,4-beta glycosidic bonds between the N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) subunits present as polymerized chains in peptidoglycan [1]. We are interested in finding out whether the fusion of SMAP-29, a small peptide shown to be able to transport a passenger protein fused to itself past the outer membrane of Gram-negative bacteria, to T4 Endolysin is able to create an antibacterial protein is that able to work against Gram-negative bacteria [2]. More importantly, the endolysin KZ144 from which Art-175 is derived is only effective against Gram-negative bacteria due to its chemotype selectivity, whereas T4 Endolysin's mechanism of action being one that targets the structural element conserved across all bacterial cell walls means that Art-E can potentially exert antimicrobial activity against an even broader spectrum of targets than Art-175 [3].

Usage

We aim to test this part for antibacterial activity by means of protein purification from host cell lysate after induction of gene expression. The antibacterial potency of this part will be compared against BBa_K1659000.

References

[1] Alhazmi, A. et al., 2014. Discovery, Modification and Production of T4 Lysozyme for Industrial and Medical Uses. International Journal of Biology, 6(4), pp.45–63. Available at: http://www.ccsenet.org/journal/index.php/ijb/article/view/38645.

[2] Briers, Y., Walmagh, M., Grymonprez, B., Biebl, M., Pirnay, J. P., Defraine, V., … Lavigne, R. 2014. Art-175 is a highly efficient antibacterial against multidrug-resistant strains and persisters of Pseudomonas aeruginosa. Antimicrobial Agents and Chemotherapy, 58(7), 3774–3784.

[3] Suvorov, M. 2008, Regulation of cell wall biosynthesis and resistance to antibiotics, University of Notre Dame. pp. 2.