Identification and characterization of the two-component cell lysis cassette encoded by temperate bacteriophage phi PYB5 of Lactobacillus fermentum

To characterize the two-component cell lysis cassette comprised of holin (Hyb5) and endolysin (Lyb5) encoded by Lactobacillus fermentum temperate bacteriophage phi PYB5, and illustrate the potential application of Lyb5 as therapeutic agents. The hyb5-lyb5 cassette was cloned from the genome library of phi PYB5, and the hyb5, lyb5 and hyb5-lyb5 cassette were expressed in E. coli BL21, respectively. The molecular weight of Hyb5 indicated by SDS-PAGE was 19 kDa, and Lyb5 was 45 kDa. Both Hyb5 and Lyb5 protein could induce cell lysis alone, resulting in the leakage of beta-galactosidase. However, the Hyb5-Lyb5 cassette lysed the host cells more rapidly and extensively. By zymogram analysis, Lyb5 exhibited a broad lytic spectrum. Overexpression of hyb5, lyb5 and hyb5-lyb5 cassette were carried out in E. coli and Lyb5 exhibited a broad lytic spectrum. The Lyb5 produced in E. coli exhibited a broad lytic spectrum against Gram-positive strains including Staphylococcus aureus as well as Gram-negative strains such as Salmonella typhi, suggesting that Lyb5 provides a potential alternative of diagnostic tools and therapeutic agents.