Identification of Three Campylobacter Lysins and Enhancement of Their Anti-Escherichia coli Efficacy Using Colicin-Based Translocation and Receptor-Binding Domain Fusion.

Abstract

The emergence of multidrug-resistant Escherichia coli, which poses a major threat to public health, has motivated the development of numerous alternative antimicrobials. Lysins are bacteriophage- and bacterium-derived peptidoglycan hydrolases that represent a new antibiotic treatment targeting bacterial cell walls. However, the bactericidal effect of native lysins on Gram-negative bacteria is restricted by the presence of an outer membrane. Here, we first evaluated the antibacterial activity of three Campylobacter-derived lysins (Clysins) against E. coli. To improve their transmembrane ability and antibacterial activities, six engineered Clysins were constructed by fusing with the translocation and receptor-binding (TRB) domains from two types of colicins (colicin A [TRBA] and colicin K [TRBK]), and their biological activities were determined. Notably, engineered lysin TRBK-Cly02 exhibited the highest bactericidal activity against the E. coli BL21 strain, with a reduction of 6.22 ± 0.34 log units of cells at a concentration of 60.1 μg/mL, and formed an observable inhibition zone even at a dose of 6.01 μg. Moreover, TRBK-Cly02 killed E. coli dose dependently and exhibited the strongest bactericidal activity at pH 6. It also exhibited potential bioactivity against multidrug-resistant E. coli clinical isolates. In summary, this study identified three lysins from Campylobacter strains against E. coli, and the enhancement of their antibacterial activities by TRB domains fusion may allow them to be developed as potential alternatives to antibiotics. IMPORTANCE Three lysins from Campylobacter, namely, Clysins, were investigated, and their antibacterial activities against E. coli were determined for the first time. To overcome the restriction of the outer membrane of Gram-negative bacteria, we combined the TRB domains of colicins with these Clysins. Moreover, we discovered that the Clysins fused with TRB domains from colicin K (TRBK) killed E. coli more effectively, and this provides a new foundation for the development of novel bioengineered lysins by employing TRBK constructs that target outer membrane receptor/transport systems. One of the designed lysins, TRBK-Cly02, exhibited potent bactericidal efficacy against E. coli strains and may be used for control of multidrug-resistant clinical isolates. The results suggest that TRBK-Cly02 can be considered a potential antibacterial agent against pathogenic E. coli.