The gp38 protein inhibits host adsorption of phage vB_EcoM_SD286

Bacteriophages (phages), which are viruses that infect bacteria, primarily use surface receptor-binding proteins (RBPs) to recognize and infect their hosts. Elucidating the function of specific RBPs is crucial for understanding phage-host interactions and developing phage-based antimicrobials.

This study characterized the Escherichia coli phage vB_EcoM_SD286, isolated from farm sewage in Shandong Province. Its morphology was observed via transmission electron microscopy. The lysis spectrum and optimal multiplicity of infection (MOI) were determined using the double-layer plate method. Stability under various pH and temperature conditions was assessed. A one-step growth curve was plotted to determine the latent period and burst size. The genome was sequenced and analyzed for open reading frames (ORFs), tRNA, virulence factors, and antibiotic resistance genes. Bioinformatic analysis suggested that the putative protein gp38 may function as an RBP. To verify this, a recombinant expression vector, pET-28a(+)-gp38, was constructed and induced in BL21(DE3) cells to produce the recombinant gp38 protein. Competitive adsorption and binding assays were conducted to evaluate its role in host recognition.

Phage vB_EcoM_SD286 exhibited an icosahedral head and a helical tail, classifying it within the Caudoviricetes class and Rosemountvirus genus. It lysed 39% of tested strains, with an optimal MOI of 0.01. The phage demonstrated stability across a broad pH range (4-12) and at temperatures below 50°C, but was completely inactivated after 20 min at 80°C. The one-step growth curve revealed a 25 min latency period and a burst size of 33 PFU/cell. Whole-genome sequencing revealed a 52,891 bp genome with 46.06% GC content, containing 74 ORFs but no tRNAs, virulence factors, or antibiotic resistance genes. The recombinant gp38 protein was successfully expressed. Subsequent competitive adsorption assays, alongside direct binding experiments between host bacteria and the gp38 protein, demonstrated that gp38 significantly inhibited phage adsorption to host bacteria.

Collectively, our findings provide preliminary evidence that gp38 is involved in the phage-host interaction of vB_EcoM_SD286, likely functioning as a receptor-binding protein. This study offers a theoretical basis for elucidating the precise bacterial receptor recognition mechanism and lays the groundwork for future development of phage-based antimicrobial agents.