Functional Selection Of Peptide That Specifically Inhibits Salmonella Typhi Adhesion To/invasion Of Human Monocytic Cells By Ribosome Display

Of the more than 2000 closely related Salmonella servers identified, Salmonella enterica serovar Typhi (S. Typhi) is an important pathogen, which infects humans exclusively and causes typhoid or enteric fever. S. Typhi bacteria efficiently invade the human internal mucosa and survive as intracellular pathogens of macrophages/monocytes and possibly dendritic cells. Recently it has been discovered that type IVB pili, encoded by the S. Typhi pil operon located in the major pathogenicity island, may be important in the pathogenesis of epidemic enteric fever, but the mechanisms between type IVB pili and monocytes is not known. To further investigate the roles of type IVB pili of S. Typhi and prepare drug that specially inhibit S. Typhi to infect monocytes, 36 random-sequence oligonucleotide and primers were designed and synthesed. A DNA random library was constructed by two rounds PCR. Then a coupled transcription/translation reaction in vitro was preceded and a ribosome display library including 12 random peptides was constructed. A 12-mer peptide (RQERSSLSKPW), binding to the structural protein prePilS of the type IVB pili of S. Typhi, was isolated with a ribosome display system. This peptide was designated as 12-mer peptide R. We found that 12-mer peptide R inhibited adhesion to/invasion of human monocytic THP-1 cells by piliated S. Typhi bacteria, but had no effects on nonpiliated S. Typhi bacteria. A random 12-mer peptide, of size and solubility equal to 12-mer peptide R, served as a control on the specificity of 12-mer peptide R. The specific interaction and binding equilibrium between the 12-mer peptide R and prePilS protein was determined by Isothermal Titration Calorimetry (ITC) and a binding constant Ka determined to be between 0.4×10~5 L. mol~-1 and 2.2×10~5 L. mol~-1. Our findings suggest that the type IV pili-binding 12-mer peptide R holds potential as an antibacterial peptide effective against S. Typhi infections, both in terms of prevention and therapeutic treatment. The data further provide insights into the understanding of the pathogenic roles of the type IVB pili of S. Typhi.