Bacteriophages possessing virion-associated depolymerasesare able to freely diffuse in the biofilm matrix. Depolymerization of bacterial exopolysaccharide results in increasing susceptibility to chemical and physical agents and to host immune defence mechanisms. Although depolymerases do not kill the bacterium, they are of special interest as anti-virulence compounds that disarm the bacterium reducing or even preventing the infection process. Anti-virulence approaches may induce less evolutionary pressure for drug resistance development in comparison to most antibacterial agents.
The genes encoding putative depolymeraseshave been identified in genomes of K.pneumoniae specific lytic phages, from the Podoviridae family (KP32 and KP34) and the Siphoviridae family (KP36). These phages have been chosen as able to form a halo zone surrounding a lysis zone when spotted on bacterial lawn. Selected enzymes have been prepared as recombinant proteins. Depolymerases originating from phages (KP36 and KP34), belonging to different families, are specific to the same capsular serotype K63 and are highly similar in the amino acid sequence in C-terminal part. The remaining KP32 depolymerase act on K20 serotype of Klebsiella capsule.
The secondary structure of these proteins have a strong β-sheet contentand is highly stable at high temperature and in wide range of pH.The oligomeric state of depolymerases is trimeric according to native PAGE electrophoresis and MALS light scattering analyses. The ability of these enzymes to degrade capsular exopolysaccharide isolated from bacterial host has been confirmed using zymography. Obtained microbiological results show high anti-virulence potential of phage encoded depolymerases against K. pneumoniae strains.