关键词: 海藻酸裂解酶, 毕赤酵母, 海藻寡糖, 诱导表达

Abstract: 【Objective】Alginate lyase was obtained efficiently, and the properties were characterized【Method】Using PCR technique, the alginate lyase gene(algL) of Pseudomonas aeruginosa was cloned and inserted into the pPIC9K expression vector and introduced into the host Pichia pastoris GS115 by PEG method. After screen, the recombinant P. pastoris strain was obtained and induced in 50mL methylotrophic culture medium. 【Results】The activity of alginate lyase reached 540U/mL and the appearance of a new protein of 40 kD detected on SDS-PAGE. It was found that the optimum temperature of the recombinant enzyme was 40°C at pH 8.5 and that activity had stabilization in 20～45℃ at pH 3.0～12.0. The activity of recombinant AlgL decreased by over 50% in the presence of 50mmol/L Zn2+, Cu2+ and Fe2+; furthermore, 50mmol/L Co2+ and Ca2+ increased it by over 50% respectively; other cations including Mn2+, Mg2+, K+ and Na+ promoted slightly the enzymatic activity in different concentration. The recombinant enzyme had effect on antibiotic sterilization as potential assistant. 【Conclusion】Recombinant alginate lyase was gained through P. pastoris expression system and analyzed, which could instruct production and application in some fields.

Key words: alginate lyase, Pichia pastoris, alga oligosaccharides, expression induced