Expression and characterization of Kunitz domain 3 and C-terminal of human tissue factor pathway inhibitor-2

• Published in: Acta biochimica et biophysica Sinica Vol. 41; no. 11; pp. 948 - 954
• Main Authors: Zhu, Lina, Wang, Jiping, Mu, Jingui, Wang, Huijun, Zhang, Chenqi, Wang, Jue, Liu, Xingang, Yan, Xiaomin, Dai, Linsen, Ma, Duan
• Format: Journal Article
• Language: English
• Published: China Oxford University Press 01.11.2009
• Subjects: Glycoproteins - chemistry; Glycoproteins - isolation & purification; Glycoproteins - metabolism; Heparin - chemistry; Humans; Pichia - genetics; Pichia - metabolism; Protein Binding; Protein Engineering - methods; Protein Structure, Tertiary; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; 丝氨酸蛋白酶抑制剂; 傅立叶变换红外光谱; 抑制物; 组织因子途径; 肝素钙; heparin; secondary structure; Kunitz domain 3 and C-terminal of hTFPI-2
• ISSN: 1672-9145; 1745-7270; 1745-7270
• DOI: 10.1093/abbs/gmp089

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 C- terminal of hTFPI-2 （hTFPI-2/KD3C）, which has the activity toward heparin calcium, have been successfully expressed in Pichia pastoris and purified by SP- Sepharose 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 α-helix and β-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 β-strands in hTFPI-2/ KD3C to different extents depending on the ratio of hTFPI-2/KD3C and heparin calcium.