Aims: The study was to examine whether glycosylation could improve the thermostability of recombinant Thermomyces lanuginosus lipase (Tll) expressed in Pichia pastoris. Methods and Results: The Tll gene was synthesized and transformed into Pichia pastoris GS115.The recombinant Tll protein was expressed and purified, and its glycosylation site was identified by LCMS/MS as Asn-33. Two nonglycosylated mutants were constructed and the variant proteins were also expressed and purified. Effects of temperature on activities of the wild-type Tll and variants were analysed. The glycosylated Tll exhibited better thermostability than nonglycosylated variants. Conclusions: Our experiments have demonstrated the improvement of Tll thermostability by Asn-33 glycosylation. Significance and Impact of the Study: This work has deepened our understanding in the mechanism of Tll thermostability and will guide us to directional improvement of lipases and even other industrial enzymes.