The present invention provides methods and compositions for production of recombinant iduronate-2-sulfatase (I2S). The invention encompasses the discovery that more potent recombinant I2S protein can be produced by cells engineered to co-express a recombinant I2S protein and a formylglycine generating enzyme (FGE). According to the invention, a cell in cell culture conditions produces I2S protein comprising at least about 70% conversion of the cysteine residue corresponding to Cys59 to Cα-formylglycine (FGly), and at a specific productivity rate of greater than about 30 picogram/cell/day and/or at a level of I2S between 0.3-fold and 10-fold higher than the level of formylglycine generating enzyme. Such cells are adaptable to various cell culture conditions, and in some embodiments are adaptable to a large-scale suspension serum-free culture. Recombinant I2S produced in accordance with the present invention can be used in enzyme replacement therapy, for example in treating Hunter syndrome.
