Identification and Characterization of a Proteinaceous Antibacterial Factor from Pseudomonas extremorientalis PEY1 Active Against Edwardsiella tarda

Pseudomonas extremorientalis PEY1, isolated from the intestinal contents of marine fish, was evaluated for the production and properties of antibacterial proteins active against Edwardsiella tarda, a major pathogen in aquaculture. Antibacterial production was maximized in a minimal medium supplemented with 1% yeast extract and 1% galactose under stationary cultivation at 25 °C and pH 7.0. Growth and bioactivity assays were conducted under varying carbon and nitrogen sources, temperatures, and pH levels. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed a distinct ~37 kDa protein band corresponding to antibacterial activity, exhibiting an inhibition zone of 2.4 ± 0.1 cm against E. tarda. The activity was completely abolished by papain digestion but remained detectable after exposure to 55 °C and pH 8, indicating that the active compound is a moderately heat-stable, proteinaceous antibacterial molecule. LC–MS/MS analysis identified the protein as a putative disulfide reductase with ~40% sequence coverage. The antibacterial factor exhibited strong physicochemical stability, retaining activity in the presence of surfactants and metal ions. Collectively, these findings demonstrate that P. extremorientalis PEY1 produces a thermostable, papain-sensitive antibacterial protein with selective activity against E. tarda, highlighting its potential as a promising natural biocontrol or postbiotic candidate for sustainable aquaculture.