Full-length and N-TIMP-3 display equal inhibitory activities toward TNF-alpha convertase.
We previously reported that tumor necrosis factor-alpha converting enzyme (TACE) was specifically inhibited by TIMP-3 but not TIMP-1, -2, and -4. Further mutagenesis studies showed that the N-terminal domain of TIMP-3 (N-TIMP-3) retained full inhibitory activity towards TACE. Full-length TIMP-3 and N-TIMP-3 exhibited indistinguishable values for the association rate constant and inhibitory affinity constant for the active catalytic domain of TACE (k(on) approximately 10(5) M(-1) s(-1) and K(app)(i) approximately 0.20 nM). Moreover, their k(on) (approximately 10(4) M(-1) s(-1)) and K(app)(i) (approximately 1.0 nM) values with a longer form of TACE (which encompasses the complete ectodomain including disintegrin, EGF and Crambin-like domains) were also shown to be similar. Detailed kinetic analyses indicated that TIMP-3 associated more quickly and with tighter final binding with TACE devoid of these C-terminal domains. We conclude that, unlike the interaction between many MMPs and TIMPs, the C-terminal domains of TIMP-3 and TACE are not essential in the formation of a tight binary complex.
Lee
M H
MH
School of Biological Sciences, University of East Anglia, Norwich NR4 7TJ, United Kingdom.
Knäuper
V
V
Becherer
J D
JD
Murphy
G
G
D051722:ADAM Proteins; D000072198:ADAM17 Protein; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics; D051379:Mice; D010446:Peptide Fragments / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics; D011480:Protease Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics; D019717:Tissue Inhibitor of Metalloproteinase-3 / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*
D017931:DNA Primers; D010446:Peptide Fragments; D011480:Protease Inhibitors; D011994:Recombinant Proteins; D019717:Tissue Inhibitor of Metalloproteinase-3; D051722:ADAM Proteins; D008666:Metalloendopeptidases; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human; C000606463:Adam17 protein, mouse