Huang W, Suzuki K, Nagase H, Arumugam S, Van Doren S R, Brew K
Department of Biochemistry and Molecular Biology, University of Miami, FL 33134, USA.
FEBS Lett. 1996 Apr 15;384(2):155-61. doi: 10.1016/0014-5793(96)00304-3.
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%.
