TiO2 nanofibers with uniform diameter about 125 nm were prepared based on sol-gel process and electrospinning technology. Protex 6L, an industrial alkaline protease, was covalently immobilized on TiO2 nanofiber through gamma-aminopropyltriethoxysilane modification and glutaraldehyde crosslinking. With 2 (v/v)% glutaraldehyde as crosslinker, the enzyme loading is about 201 mg (g nanofiber membrane)(-1), and the specific activity of the immobilized Protex 6L is 2.45 mu mol h(-1) ml(-1) mg(-1) protein for synthesis of sucrose monolaurate from sucrose and vinyl laurate. The optimal condition for sucrose monolaurate production is 5% (v/v) water content in DMSO/2-methyl-2-butanol solvent mixture and 50A degrees C. Under this condition, 97% conversion was achieved within 36 h by nanofibrous Protex 6L, which is corresponding to a productivity 34 times higher than that of most widely used Novozym 435. After 10 cycles reuse, nanofibrous Protex 6L retained 52.4% of its original activity.