ACTIVATION OF PROGELATINASE-B (PROMMP-9) BY ACTIVE COLLAGENASE-3 (MMP-13)

Human progelatinase B was activated by collagenase-3 in a time-depende nt fashion, Activation proceeded through an intermediate form of M-r 8 6 000 to the final active form of M-r 82 000. N-terminal amino acid se quence determination demonstrated that the Glu40-Met41 peptide bond wa s initially hydrolysed followed by cleavage of the Arg87-Phe88 peptide bond releasing the rest of the propeptide domain which was accompanie d by the achievement of maximal enzymatic activity as revealed using a quenched fluorescent substrate. Kinetic analysis of activation reveal ed that the rates were dependent on the concentration of the proenzyme as well as active collagenase-3. Active gelatinase B did not contribu te to the activation rate of the proenzyme initiated by collagenase-3 and our results indicate that progelatinase B activation proceeds via bimolecular cleavage with collagenase-3 involving sequential cleavage of the propeptide in two steps. The activation rates were not dependen t on C-terminal domain interactions between progelatinase B and collag enase-3, as assessed using wild-type and C-terminal deletion mutants o f both enzymes. Since elevated levels of both gelatinase B and collage nase-3 have been observed in arthritis and breast cancer pathology the se enzymes may well form a proteolytic cascade in these diseases which allows rapid turnover of the extracellular matrix.