Expression and characterization of the first kunitz domain of human tissue factor pathway inhibitor-2.
Human tissue factor pathway inhibitor-2 (hTFPI-2) has three kunitz domains whose structure and function are unclear. We expressed the first kunitz domain of hTFPI-2 (hTFPI-2/KD1) as functional form using Pichia pastoris and investigated its characterization. In the experiment, hTFPI-2/KD1 can inhibit the plasmin and trypsin activity and the Ki of hTFPI-2/KD1 towards plasmin (30nM) and trypsin (50nM) was determined as 10 and 7nM by chromogenic assay, respectively. hTFPI-2/KD1 can also inhibit MMP-2 and MMP-9 in zymography assay. Furthermore, the inhibition of hTFPI-2/KD1 to the Matrigel invasion by HT-1080 is also described. This study provides a method to produce hTFPI-2/KD1 efficiently and some insights into the structure and function of hTFPI-2/KD1.
Kong
Desheng
D
Key Laboratory of Molecular Medicine, Ministry of Education, Dong'an Road 130#, Fudan University, Shanghai 200032, China.
Ma
Duan
D
Bai
Hao
H
Guo
Hongshen
H
Cai
Xu
X
Mo
Wei
W
Tang
Qiqun
Q
Song
Houyan
H
D045744:Cell Line, Tumor; D005341:Fibrinolysin / Q000378:metabolism; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020782:Matrix Metalloproteinases / Q000378:metabolism; D009361:Neoplasm Invasiveness; D010843:Pichia / Q000235:genetics; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000032:analysis / Q000302:isolation & purification / Q000494:pharmacology; D014357:Trypsin / Q000378:metabolism
D006023:Glycoproteins; D011994:Recombinant Proteins; C021240:tissue-factor-pathway inhibitor 2; D014357:Trypsin; D005341:Fibrinolysin; D020782:Matrix Metalloproteinases