Cloning, characterization and functional expression of a new beta-D-fructofuranosidase (Os beta fruct2) cDNA from Oryza sativa.

A new cDNA (Os beta fruct2) encoding an acid beta-D-fructofuranosidase from rice has been cloned, sequenced and expressed in Pichia pastoris. The full-length cDNA is 2453 base pairs long and encodes a pre-pro-protein of 682 amino acids. The cDNA fragment coding for mature enzyme was sub-cloned into vector pPICZalphaA for extracellular expression in the methylotrophic yeast. The recombinant product was purified by Ni2+-nitrilotriacetic acid agarose column and biochemically characterized. The enzyme could hydrolyze sucrose and raffinose. Molecular mass is 66 kDa. The activity optimum was at pH 4.8 and 40 degrees C.