Isolation and purification of an effective thermal stable alginate lyase on hospital Pseudomonas aeruginosa alginate

The bacterial alginate lyase is a periplasmic enzyme that is important in the production and disruption of alginate. Characterization of this enzyme such as molecular weight, pI, thermal stability, optimum pH and temperature, and effectiveness on bacterial alginate can differ in the strains of a bacterium. In this study, to obtain an alginate lyase containing specific and efficient properties in destroying of bacterial alginate, a mucoid strain of Pseudomonas aeruginosa isolated from patient‘s sputum was used. Since alginate lyase also has a role in alginate production, this strain was selected for this assay. Alginate lyase was extracted from periplasmic space by heat shock method and was purified by using suitable concentration of ammonium sulfate as well anionic exchange chromatography. Subsequently, enzyme activity was assayed by using thiobarbitoric acid method. The relative molecular weight of the enzyme was estimated about 60 kDa. The effect of reaction time and pH, substrate concentration and impact of temperature on alginate lyase activity was surveyed. Results showed that the optimum activity of the enzyme was achieved at 0.4 mg/ml substrate concentration after 20 min in 37˚C and pH 8.5. The enzymatic residual activity after 4 hours incubation in 80˚C was determined 31.33% that indicated to the spectacular thermal stability of the enzyme.This enzyme has the ability to disrupt the alginate of P. aeruginosa M 8821, and this ability associated with the thermal stability make it an excellent option for adjunctive therapy in the combat against antibiotic-resistant of P. aeruginosa infections.