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Interaction between tissue inhibitor of metalloproteinases-2 and progelatinase A: immunoreactivity analyses.

ABSTRACT: By immunoreactivity analysis using monoclonal antibodies, we showed that the C-terminal domain [R415-631; R is residue] of progelatinase A [pro-matrix metalloproteinase-2 (proMMP-2); EC 3.4.24.24] affected the immunoreactivity of a one-step sandwich enzyme immunoassay (sandwich EIA) for tissue inhibitor of metalloproteinases-2 (TIMP-2) in exactly the same way as does proMMP-2 [Fujimoto, Zhang, Iwata, Shinya, Okada and Hayakawa (1993) Clin. Chim. Acta 220, 31-45], confirming that the C-terminal domain ("tail" portion of TIMP-2 participates in the binding with the C-terminal domain of proMMP-2. We also demonstrated that not only the C-terminal domain but also the N-terminal domain (R1-417) of proMMP-2 bound to TIMP-2 in a 1:1 molar ratio. The binding of each individual domain to TIMP-2, however, was weak enough that either domain could be fully replaced by proMMP-2 through the same binding sites as does proMMP-2, and also that the high-order structure of proMMP-2 allows a more stable binding to TIMP-2. We further confirmed that TIMP-2 complexed with the N-terminal domain of pro-MMP-2 had fully inhibitory activity against the collagenolytic activity of MMP-1. We also demonstrated that either the interstitial collagenase-TIMP-2 complex or the gelatinase B(MMP-9)-TIMP-2 complex was able to form a ternary complex with proMMP-2 in a 1:1 molar ratio, clearly indicating that there are two distinct binding sites, one specific for proMMP-2 complex, but the binding seemed to be less stable than the binding with TIMP-2 alone. Even in the presence of a 10-fold molar excess of the N-terminal domain, ternary complex formation was not observed between the N-terminal domain and the MMP-9--TIMP-2 complex. These clear differences might be ascribed to some significant conformational change(s) evoked in the TIMP-2 molecule, or hindrance of a part of the N-terminal domain binding site of TIMP-2 by complex formation with MMP-9.

SUBMITTER: Fujimoto N

PROVIDER: S-EPMC1216985 | biostudies-other | 1996 Feb

REPOSITORIES: biostudies-other

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Project description:BackgroundMatrix metalloproteinases (MMPs) are involved in remodeling of the extracellular matrix (ECM) during pregnancy and parturition. Aberrant ECM degradation by MMPs or an imbalance between MMPs and their tissue inhibitors (TIMPs) have been implicated in the pathogenesis of preterm labor, however few studies have investigated MMPs or TIMPs in maternal serum. Therefore, the purpose of this study was to determine serum concentrations of MMP-3, MMP-9 and all four TIMPs as well as MMP:TIMP ratios during term and preterm labor.MethodsA case control study with 166 singleton pregnancies, divided into four groups: (1) women with preterm birth, delivering before 34 weeks (PTB); (2) gestational age (GA) matched controls, not in preterm labor; (3) women at term in labor and (4) at term not in labor. MMP and TIMP concentrations were measured using Luminex technology.ResultsMMP-9 and TIMP-4 concentrations were higher in women with PTB vs. GA matched controls (resp. p = 0.01 and p<0.001). An increase in MMP-9:TIMP-1 and MMP-9:TIMP-2 ratio was observed in women with PTB compared to GA matched controls (resp. p = 0.02 and p<0.001) as well as compared to women at term in labor (resp. p = 0.006 and p<0.001). Multiple regression results with groups recoded as three key covariates showed significantly higher MMP-9 concentrations, higher MMP-9:TIMP-1 and MMP-9:TIMP-2 ratios and lower TIMP-1 and -2 concentrations for preterm labor. Significantly higher MMP-9 and TIMP-4 concentrations and MMP-9:TIMP-2 ratios were observed for labor.ConclusionsSerum MMP-9:TIMP-1 and MMP-9:TIMP-2 balances are tilting in favor of gelatinolysis during preterm labor. TIMP-1 and -2 concentrations were lower in preterm gestation, irrespective of labor, while TIMP-4 concentrations were raised in labor. These observations suggest that aberrant serum expression of MMP:TIMP ratios and TIMPs reflect pregnancy and labor status, providing a far less invasive method to determine enzymes essential in ECM remodeling during pregnancy and parturition.

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Interaction between tissue inhibitor of metalloproteinases-2 and progelatinase A: immunoreactivity analyses.

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