One-step purification and characterization of alginate lyase from clinical with destructive activity on bacterial biofilm.

OBJECTIVES

is a Gram-negative and aerobic rod bacterium that displays mucoid and non-mucoid phenotype. Mucoid strains secrete alginate, which is the main agent of biofilms in chronic infections, show high resistance to antibiotics; consequently, the biological disruption of mucoid biofilms is an attractive area of study for researchers. Alginate lyase gene is a member of alginate producing operon which by glycosidase activity produces primer for other enzymes in this cluster. Also this activity can destroy the extracellular alginate; therefore this enzyme participates in alginate production and destruction pathway. Alginate lyase causes detachment of a biofilm by reducing its adhesion to the surfaces, and increases phagocytosis and antibiotic susceptibility. In this study, alginate lyase was purified in just one step and its properties were investigated.

MATERIALS AND METHODS

The purification was done by affinity chromatography, analysed by SDS-PAGE, and its effect on biofilms was surveyed by micro titer plate assay and SEM. The substrate specificity of the enzyme was determined by PCR.

RESULTS

Alginate lyase from isolate 48 was purified in one step. It is more thermally resistant than alginate lyase from PAO1 and poly M, poly G and poly MG alginate were the substrate of this enzyme. Moreover, it has an eradication effect on biofilms from 48 and PAO1.

CONCLUSION

In this study an alginate lyase with many characteristics suitable in medicine such as thermal stability, effective on poly M alginate, and bacterial biofilm destructive was introduced and purified.