Activation of progelatinase A and progelatinase A TIMP-2 complex by membrane type 2 matrix metalloproteinase

作者：Kolkenbrock H^*; HeckerKia A; Orgel D; Ulbrich N; Will H

来源：Biological Chemistry, 1997, 378(2): 71-76.

DOI：10.1515/bchm.1997.378.2.71

• progelatinase A
• TIM P-2
• membrane-type-MMP

C-terminal truncated membrane-type 2 matrix metalloproteinase (MT2-MMP(1-269)), comprising prodomain and catalytic domain, was expressed as a soluble protein in Escherichia coli, Unlike the corresponding form of MT1-MMP, which can be isolated as a 31 kDa protein, MT2-MMP(1-269) proved to be comparatively instable, and already the freshly isolated preparation displayed several proteins in SDS-PAGE representing MT2-MMP(1-269) (33 kDa) and four N-truncated forms with N-termini methionine(32) (30 kDa), isoleucine(37) (30 kDa), leucine(84) (24 kDa), and leucine(93) (22 kDa), the catalytic domain, After thawing of frozen preparations the 33 and the 30 kDa proforms were no longer detectable in SDS-PAGE, and only the 24 and 22 kDa forms remained.
The catalytic domain of MT2-MMP activated progelatinase A as well as the progelatinase A/TIMP-2 complex by cleaving the 72 kDa progelatinase A to yield 67 kDa gelatinase A, which is then transformed into 62 kDa gelatinase A, The 62 kDa form is about twice as active as the 67 kDa form towards the synthetic substrate N-(2,4)-dinitrophenyl-Pro-Gln-Gly-Ile-Ala-Gly-Gln-D-Arg, No significant difference in activity was found between free and complexed gelatinase A forms, The activation of the progelatinase A/TIMP-2 complex proceeds in two steps: At first MT2-MMP is inhibited by the progelatinase A/TIMP-2 complex, whereby a ternary complex, progelatinase A/TIMP-2/MT2-MMP, is generated, This ternary complex is then activated by excess MT2-MMP, Our results suggest a mechanism for spatially regulated extracellular gelatinase A activity mediated by activation with membrane-type MMPs: Free gelatinase A is released into the extracellular space, while gelatinase A/TIMP-2 bound to MT-MMP remains anchored on the cell surface.

• 出版日期1997-2