Endolysin (gp1.2) from the Paenibacillus polymyxa CCM 7400 temperate phage phiBP has a modular structure consisting of an N-terminal region with a catalytic glycosyl hydrolase 25 domain and a C-terminal cell wall-binding domain. Another studied endolysin is from the Brevibacterium flavum CCM 251 lytic phage BFK20. The endolysin BFK20 (gp24´) has also a modular structure consisting of an N-terminal amidase_2 domain (gp24CD) and a C-terminal cell wall binding domain (gp24BD). Previously, the entire genes of both endolysins and corresponding fragments containing catalytic domain and binding domains separately were cloned into expression vector pET28a+ [1, 2]. The presence of a cell wall-binding domain in endolysin phiBP was found to be essential, as the phiBP endolysin was fully active only as a full-length protein. On the contrary the BFK20 catalytic domain activity is clearly inhibited by the presence of the cell wall binding domain. In our search for novel antimicrobial agents against Paenibacillus larvae and corynebacteria, we constructed chimeric lysin named as BP-H1 by fusing the catalytic domain of endolysin phiBP with the cell wall binding domain of endolysin BFK20 and two chimeric lysins named as BFK-H2-1 and BFK-H2-2 by fusing the catalytic domain of endolysin BFK20 with shorter and longer region of the cell wall binding domain of endolysin phiBP. The recombinant proteins were expressed in Escherichia coli and purified by affinity chromatography. The lytic activities of chimeric endolysins were tested on cell wall substrates from paenibacilli, bacilli and corynebacteria and the antimicrobial activity against Paenibacillus larvae was studied as well. Our results showed that chimeric lysin BFK-H2-1 is fully active towards paenibacilli, bacilli and corynebacteria.

Acknowledgement: This work was supported by the VEGA Grant 2/0122/14 and APVV-0098-10.

Reference:
[1]. Gerova et al. (2011) FEMS Microbiol. Lett. 321(2): 83-91
[2]. Ugorcakova et al. (2015) FEMS Microbiol. Lett. 362(fnv098): 1-9