Folding and characterization of the amino-terminal domain of human tissue inhibitor of metalloproteinases-1 (TIMP-1) expressed at high yield in E. coli

FEBS Lett. 1996 Apr 15;384(2):155-61. doi: 10.1016/0014-5793(96)00304-3.

Abstract

Methods are described for producing an active amino-terminal domain of tissue inhibitor of metalloproteinases-1 (N-TIMP-1) from inactive protein expressed as inclusion bodies in E. coli. Yields exceed 20 mg per litre of bacterial culture. Activity measurements, CD spectroscopy and NMR spectroscopy of the 15N-labeled protein show that it is fully active, homogeneous in conformation and suitable for high-resolution structural analysis. The affinity of N-TIMP-1 for matrix metalloproteinases 1, 2 and 3 is 6-8-fold less than that of the recombinant full-length protein, indicating that deletion of the C-terminal domain reduces the free energy of interaction by < 10%.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • CHO Cells
  • Circular Dichroism
  • Cricetinae
  • Escherichia coli / metabolism*
  • Glycoproteins / biosynthesis*
  • Glycoproteins / chemistry
  • Glycoproteins / genetics
  • Humans
  • Magnetic Resonance Spectroscopy
  • Molecular Sequence Data
  • Promoter Regions, Genetic
  • Protein Conformation
  • Protein Folding
  • Recombinant Fusion Proteins / biosynthesis*
  • Recombinant Fusion Proteins / chemistry
  • Thermodynamics
  • Tissue Inhibitor of Metalloproteinases

Substances

  • Glycoproteins
  • Recombinant Fusion Proteins
  • Tissue Inhibitor of Metalloproteinases