Part:BBa_K3416032

Introduction

Vilnius-Lithuania iGEM 2020 project FlavoFlowincludes three goals towards looking for Flavobacterium disease-related problems’ solutions. The project includes creating a rapid detection kit, based on HDA and LFA, developing an implement for treating a disease, and introducing the foundation of edible vaccines. This part was used for the second goal- treatment - of the project FlavoFlow.

Biology

Bacteriophage lysins are enzymes encoded by bacteriophages that cut peptidoglycans in bacterial cell walls(Fig.1). Endolysins are missing the signal peptide sequence, that is why they are not transferred through the cytoplasmic membrane1 . These enzymes have a high rate of lytic action, they also can work upon antibiotic-resistant bacterial strains as well as their ability to degrade bacterial biofilms. LysECD7 is an endolysin of the putative zinc D-alanyl-D-alanine carboxypeptidase (peptidase M15 family) of the lytic bacteriophage ECD72. This bacteriophage is related to the Myoviridae family(Fig. 2) and infects the Escherichia species³. Normally, endolysins act from within the bacterial host at the end of the bacteriophage’s replicative cycle, allowing the release of viral offspring.

Figure 1. Endolysin activity mechanism⁵. a) Endolysin can not access peptidoglycan. b) Endolysins are able penetrate the outer membrane because: 1) they have specific membrane passing abilities; 2) they work together with permeabilizing reagents to enter through the outer membrane; 3) they are fused with self-promoted uptake mediators.

LysECD7 might become a potential therapeutic agent for antimicrobial drugs, since results revealed a broad spectrum of activity against Gram-negative bacteria^4,5. However, this endolysin action depends on a lot of aspects such as buffer conditions, salts concentrations, the pH levels of the medium. For the most efficient use of endolysins, it is important that they maintain their activity under these conditions: high salt composition and pH from 5.5 to 7.42.

Figure 2. Typical structure of a myovirus³.

References

1. Jarábková, V., Tišáková, L. & Godány, A. Phage Endolysin: A Way To Understand A Binding Function Of C-Terminal Domains, A Mini Review. Nova Biotechnologica et Chimica 14, 117–134 (2015).
2. Antonova, N. et al. Broad Bactericidal Activity of the Myoviridae Bacteriophage Lysins LysAm24, LysECD7, and LysSi3 against Gram-Negative ESKAPE Pathogens. Viruses 11, 284 (2019).
3. Myoviridae. Virus Taxonomy 46–62 (Elsevier, 2012).
4. Antonova, N. P. et al. Modulation of Endolysin LysECD7 Bactericidal Activity by Different Peptide Tag Fusion. Biomolecules 10, 440 (2020).
5. Schmelcher, M. & Loessner, M. J. Bacteriophage endolysins: applications for food safety. Current Opinion in Biotechnology 37, 76–87 (2016).