Abstract:

Streptomyces diastatochromogenes TUST2, which was isolated from Hainan province, produced the antimicrobial poly(amino acid), ε-poly-L-lysine. In this study, the ε-poly-L-lysine-degrading enzyme was purified from this strain and its properties of this enzyme were determined. Preliminary data suggested that the ε-poly-L-lysine-degrading enzyme was a cell membrane associated protein. To extract this enzyme, bacterial cells were collected and disrupted with an ultrasonic oscillator, the membrane fraction were solubilized with 1.0 mol/L NaSCN solution. The coarse enzyme extraction was subjected to Sephadex G100 column for purification. With 100 mmol/L phosphate buffer as elution solution, the fractions with the enzyme activity were collected. The purified sample was analyzed with SDS-PAGE and the subunit molecular mass of the enzyme was estimated to be about 54700. The enzyme was stable between pH 6.0 and 9.0, with a maximum at pH=7.0. The optimum temperature was 30 ℃, and no significant activity loss was observed when the enzyme was incubated at 10—50 ℃ for 30 min. The effect of different metal ions on the activity of the enzyme were also investigated, some metal ions, including Zn²⁺, Fe³⁺, and Cu²⁺, could increase the enzyme activities by 29.72%, 15.85%, and 15.08%, respectively; while some other metal ions, including Ag⁺, Hg²⁺, Co²⁺ and Mn²⁺, strongly inhibited the enzyme activity. The enzyme activity was not affected by Ca²⁺, K⁺ and Ba²⁺. Experiment results also showed that the enzyme activity increased 10% by addition of 4％Tween-80, while EDTA strongly inhibited the enzyme activity. These results suggested that the specificities of this enzyme are similar to that of the ε-poly-L-lysine-degrading enzyme from streptomyces albulus.

Key words: Streptomyces diastatochromogenes; ε-Poly-L-lysine -degrading enzyme; Purification and characte-rization