Tyrosine 36 plays a critical role in the interaction of the AB loop of tissue inhibitor of metalloproteinases-2 with matrix metalloproteinase-14.
The tissue inhibitor of metalloproteinases-2 (TIMP-2) is potentially an important inhibitor of all known matrix metalloproteinases (MMPs). However, it has been shown to undergo specific interactions with both MMP-2 (gelatinase A) and MMP-14 (MT1-MMP), and it has been proposed that these three proteins function as a cell surface-based activation cascade for matrix metalloproteinases and as a focus of proteolytic activity. In this study, we have carried out mutagenesis and kinetic analyses to examine the unique interactions between the AB loop of TIMP-2 and MMP-14. The results demonstrate that the major binding contribution of the AB loop is due solely to residue Tyr-36 at the tip of the hairpin. From this work, we propose that TIMP-2 may be engineered to abrogate MMP-14 binding, whereas its binding properties for other MMPs, including MMP-2, are maintained. Mutants of TIMP-2 with more directed specificity may be of use in gene therapeutic approaches to human disease.
Williamson
R A
RA
School of Biological Sciences, University of East Anglia, Norwich NR4 7TJ, United Kingdom.
Hutton
M
M
Vogt
G
G
Rapti
M
M
Knäuper
V
V
Carr
M D
MD
Murphy
G
G
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D018360:Crystallography, X-Ray; D006801:Humans; D007700:Kinetics; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000737:chemistry* / Q000378:metabolism*; D014443:Tyrosine
D011994:Recombinant Proteins; D019716:Tissue Inhibitor of Metalloproteinase-2; D014443:Tyrosine; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases