Inulinase is an important enzyme used in the high fructose syrup and other related industries. A more cost-effective approach is required for producing highly active inulinase. In this study, the gene encoding inulinase of the yeast Kluyveromyces marxianus CBS 6556 was expressed in methylotrophic host Pichia pastoris and secretory production of recombinant inulinase (rKmINU) in the yeast under methanol induction was achieved. The purified rKmINU showed a specific activity of 2714U/mg, which is over 12-fold higher than those of other inulinases described previously. It displayed excellent stability from 30 to 50°C and pH 3.0–5.0, and the half-life of rKmINU was over 96h under these conditions. Moreover, rKmINU saccharified Jerusalem artichoke tuber juice effectively.