Synthesis and refolding of human TIMP‐2 from E. coli, with specific activity for MMP‐2

FEBS Letters

Garbisa, Spiridione²  Masiero, Laura²  Onisto, Maurizio²  Negro, Alessandro¹ 

[1] Department of Biochemistry, School of Veterinary Medicine, University of Padova, Via Trieste 75, 35121 Padova, Italy;Institute of Histology and General Embryology, University of Padova, Via Trieste 75, 35121 Padova, Italy

关键词: Tissue inhibitor of metalloproteinase;    Type IV collagenase;    Protein refolding;    Ion metal affinity column;   

DOI  :  10.1016/0014-5793(95)00073-I

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Tissue inhibitors of metalloproteinase (TIMPs) are inhibitory counterparts of collagenases, containing 12 cysteine residues paired to six internal disulphide bridges. TIMP-2, an inhibitory protein of 72 kDa gelatinase/type IV collagenase (MMP-2), was expressed in Escherichia coli as a fusion protein with a 34 amino acid NH₂-linked tail containing six consecutive histidine residues. The protein was purified in a single-step using an ion metal affinity column (IMAC) in denaturing conditions. The immobilized fusion TIMP-2 protein was refolded at a high concentration in the column, producing about 5 mg of protein per litre of bacterial cells. It shows specific binding and inhibitory activity against MMP-2, but has no effect against 92 and 45 kDa gelatinases.

【 授权许可】

Unknown