Biochemical characterization of the catalytic domain of membrane-type 4 matrix metalloproteinase

A C-terminal truncated form of membrane-type 4 matrix metalloproteinase (MT 4-MMP; MMP 17), lacking the hemopexin-like and transmembrane domain, was ex pressed in Escherichia coli. The catalytic domain was produced by tryptic a ctivation of the recombinant proenzyme and proved to be catalytically activ e towards the fluorogenic substrate for matrix metalloproteinases (7-methox ycoumarin-4-yl) acetyl,Pro-Leu-Gly-Leu(3- (2,4-dinitrophenyl)-L-2,3-diamino lprapionyl)-Ala-Arg-NH2. In contrast to the other three MT-MMPs (MT1-, MT2-, and MT3-MMP), the catal ytic domain of MT4-MMB does not activate progelatinase A, nor does it hydro lyze one of the offered extracellular matrix (ECM) proteins, such as collag en types I, II, III, IV, and V, gelatin, fibronectin, laminin or decorin. T IMP-1, a poor inhibitor of MT1-, MT2- and MT3-MMP, suppresses MT4-MMP activ ity effectively. The progelatinase A/TIMP-2 complex that usually reacts lik e TIMP-2 also inhibits MT4-MMP. TIMP-2, a strong inhibitor of other MT-MMPS , inhibits MT4-MMP at low concentrations. With increasing TIMP-2 concentrat ion, however, activity passes through a minimum and then increases until at high TIMP-2 concentration the activity is the same as in the absence of TI MP-2. TIMP-1 or the progelatinase A/TIMP-2 complex do not prevent reactivat ion of MT4-MMP catalytic domain at high TIMP-2 concentrations.