Synthesis and Preliminary Characterization of the Antimicrobial Peptide P-113 Using SPPS

PRESENTED BY: JUNE CHOI, WILEY WINKLER, BRENDEN, ALLEN BRYAN Title: Synthesis and Preliminary Characterization of the Antimicrobial Peptide P-113 Using SPPS Introduction: Peptides are short chains of amino acids that play diverse and vital roles within the human body. As the fundamental units that make up proteins, they play a crucial role in the growth, repair, maintenance, and even defense of cells and our body. The focus of the research was on the synthesis of the fragments of the P-113 antimicrobial peptide (AKRHHGYKRKFH). Found in human saliva,it exhibits activity against various oral pathogens. Through the selection of this peptide sequence, we sought to investigate how easily the peptide fragments (AKRH, HGYK, and RKFH) could be detected with mass spectrometry, as well as set up future experiments for testing the antimicrobial efficacy of these fragments against a range of bacterial strains. Method: Solid Phase Peptide Synthesis (SPPS) was employed to construct these peptides. In this technique, the C-terminal amino acid is directly attached to resin beads, allowing for the controlled, stepwise chain elongation. Each amino acid has an Fmoc (9-fluorenylmethyloxycarbonyl) protecting group on the 𝞪-amino group, which prevents unwanted side reactions and ensures selective coupling. The deprotection step with 20% Piperidine in DMF removed this protecting group, and the coupling steps added the amino acid to the peptide. This process continues sequentially from the C-terminus to the N-terminus. Once synthesis was complete, the peptide was cleaved from the resin, releasing it into solution. The crude product was then purified to remove excess reagents and byproducts, and subsequently freeze-dried with a lyophilizer. Finally, the percent yield of the peptide was calculated. Result: By following this procedure, the desired antimicrobial peptide fragments were successfully synthesized and their purity was evaluated by mass spectrometry. The peptide sequence will be used for future studies on its properties and antimicrobial efficacy by using mass spectrometry methods and zone inhibition experiments.