Double-trefoil domain mutant of human trefoil factor 3 (hTFF3) was constructed through gene engineering and inserted into to pPIC9K plasmid, then expressed in the methylotrophic yeast Pichia pastoris. The recombinant protein was purified through S-Sepharose, Q-Sepharose and Sephacryl S-100. Molecular weight of the expressed protein was about 12 000 identified by SDS-PAGE and Western-blotting. N-terminal amino acid analyses proved that the expressed hTFFE3 dimeric form mutant was identical to the native N-terminus. Two protein peaks corresponding to the monomeric and dimeric form of Di-hTFF3 mutant were identified by the mass spectrometry. The boiactivity of the mutant was the same as the native dimeric form of hTFF3, and higher than the of the monomeric mutant.

Key words: MUTANT, VITAL, ESCHERICHIA COLI, YEAST, PROTEIN