Expression and characterization of Kunitz domain 3 and C-terminal of human tissue factor pathway inhibitor-2.
Human tissue factor pathway inhibitor-2 (hTFPI-2) is a serine protease inhibitor and its inhibitory activity is enhanced by heparin. The Kunitz domain 3 and Cterminal of hTFPI-2 (hTFPI-2/KD3C), which has the activity toward heparin calcium, have been successfully expressed in Pichia pastoris and purified by SPSepharose and heparin-Sepharose chromatography. The Fourier transformed infrared spectroscopy (FTIR), Raman spectroscopy, and circular dichroism (CD) experiment results implied that hTFPI-2/KD3C contained small contents of alpha-helix and beta-strand, but large amounts of random coil and two kinds of disulfide bonds, gauche-gauche-gauche (ggg) and trans-gauchetrans (tgt). The interaction of hTFPI-2/KD3C with heparin calcium was investigated by CD. It was found that heparin calcium induced b-strands in hTFPI-2/ KD3C to different extents depending on the ratio of hTFPI-2/KD3C and heparin calcium.
Zhu
Lina
L
Center of Analysis and Measurement, Fudan University, Shanghai 200433, China.
Wang
Jiping
J
Mu
Jingui
J
Wang
Huijun
H
Zhang
Chenqi
C
Wang
Jue
J
Liu
Xingang
X
Yan
Xiaomin
X
Dai
Linsen
L
Ma
Duan
D
D006023:Glycoproteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D006493:Heparin / Q000737:chemistry*; D006801:Humans; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D006023:Glycoproteins; D011994:Recombinant Proteins; C021240:tissue-factor-pathway inhibitor 2; D006493:Heparin