Preparation and Antimicrobial Function Research of a Defensin Polypeptides SPDef1 from <i>Scorpiops_pococki in Tibet</i>

Objective: This study was designed to produce the defensin peptide SPDef1 (<i>Scorpiops pocoki defensin </i>1) via the air oxidation method, evaluate its antibacterial activity, and preliminarily investigate its antibacterial mechanism. Methods: SPDef1-O was obtained through the air oxidation method. Molecular weight and structural characterization of the prepared peptides using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF/MS) and circular dichroism spectroscopy. The minimum inhibitory concentration (MIC) of SPDef4 was determined using the broth dilution method. The antibacterial mechanism was preliminarily explored using DMAO/PI dual staining in conjunction with confocal microscopy and Transmission Electron Microscopy (TEM). Results: SPDef1-O prepared via the air oxidation method exhibits significant antibacterial activity against the Gram-positive bacterium Staphylococcus aureus strain AB94004, MIC as low as 0.25 μM, also exhibits moderate antibacterial activity against the Gram-negative bacterium Escherichia coli ATCC 25922 (MIC = 4 μM). DMAO/PI dual staining and TEM revealed that SPDef1-O disrupts cell membrane integrity to exert its antibacterial effect. Discussion: The defensin SPDef1 emerges as a novel member of scorpion-derived antimicrobial peptides with notable antibacterial activity. Future investigations should expand its antibacterial spectrum assessment and explore its potential as a broad-spectrum antimicrobial agent.