Partial Purification and Biochemical Characterization of L-lysine α-Oxidase from Clinical Isolates of Pseudomonas aeruginosa

Background: L-lysine α-oxidase (LO) is an oxidative enzyme with notable therapeutic and industrial potential due to its capacity to degrade lysine and generate cytotoxic by-productssuch as hydrogen peroxide. Pseudomonas aeruginosa is a known microbial source of LO, and itsclinical prevalence in nosocomial infections supports its exploration for enzyme production.Methodology: A total of 130 clinical samples were collected from various infection sites inBaghdad hospitals between October and December 2024. LO production was screened, and themost productive isolate (P6) was selected for purification and characterization. Purificationinvolved ammonium sulfate precipitation, DEAE-cellulose ion-exchange chromatography, andSephadex G-150 gel filtration. The enzyme was further characterized by SDS-PAGE, and itsactivity and stability were tested under varying pH, temperature, metal ions, and inhibitorconditions.Results: A total of 130 clinical samples were collected from different infection sites, including wounds, burns, urine, and sputum. Out of these, 20 isolates (15%) were identified asPseudomonas aeruginosa. The initial identification was based on colony morphology on cetrimide agar, characteristic pigmentation, and Gram staining. Further confirmation was achievedthrough standard biochemical tests (oxidase positive, growth at 42 °C, etc.) and automatedidentification using the VITEK 2 system. The specific activity of LO reached 80 U/mg proteinafter purification, with a purification fold of 10 and a yield of 49.5%. SDS-PAGE estimated themolecular weight of LO to be 54 kDa. Optimal enzymatic activity and stability were observedat pH 7 and 37 °C. Calcium and potassium ions enhanced enzyme activity, while mercury ionsshowed strong inhibition. The enzyme retained high activity in the presence of EDTA at lowconcentrations.Conclusion: The successful isolation and purification of L-lysine α-oxidase from a local P.aeruginosa isolate demonstrate its potential for therapeutic and industrial applications. Itsoptimal activity at physiological conditions and tolerance to certain ions further support itsapplicability. Future studies should focus on scaling production and testing cytotoxicity againstcancer cell lines