Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.23 (MMP)
 4,246 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Membrane type 1 matrix metalloproteinase (MT1-MMP) is a type I transmembrane MMP shown to play a critical role in normal development and in malignant processes. Emerging evidence indicates that MT1-MMP is regulated by a process of ectodomain shedding. Active MT1-MMP undergoes autocatalytic processing on the cell surface, leading to the formation of an inactive 44-kDa fragment and release of the entire catalytic domain. Analysis of the released MT1-MMP forms in various cell types revealed a complex pattern of shedding involving two major fragments of 50 and 18 kDa and two minor species of 56 and 31-35 kDa. Protease inhibitor studies and a catalytically inactive MT1-MMP mutant revealed both autocatalytic (18 kDa) and non-autocatalytic (56, 50, and 31-35 kDa) shedding mechanisms. Purification and sequencing of the 18-kDa fragment indicated that it extends from Tyr(112) to Ala(255). Structural and sequencing data indicate that shedding of the 18-kDa fragment is initiated at the Gly(284)-Gly(285) site, followed by cleavage between the conserved Ala(255) and Ile(256) residues near the conserved methionine turn, a structural feature of the catalytic domain of all MMPs. Consistently, a recombinant 18-kDa fragment had no catalytic activity and did not bind TIMP-2. Thus, autocatalytic shedding evolved as a specific mechanism to terminate MT1-MMP activity on the cell surface by disrupting enzyme integrity at a vital structural site. In contrast, functional data suggest that the non-autocatalytic shedding generates soluble active MT1-MMP species capable of binding TIMP-2. These studies suggest that ectodomain shedding regulates the pericellular and extracellular activities of MT1-MMP through a delicate balance of active and inactive enzyme-soluble fragments.
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PMID:Complex pattern of membrane type 1 matrix metalloproteinase shedding. Regulation by autocatalytic cells surface inactivation of active enzyme. 1200 57

An understanding of the regulatory mechanisms that control the activity of membrane type-1 matrix metalloproteinase (MT1-MMP), a key proteinase in tumor cell invasion, is essential for the design of potent and safe anti-cancer therapies. A unique proteolytic pathway regulates MT1-MMP at cancer cell surfaces. The abundance of proteolytic enzymes in cancer cells makes it difficult to identify the autocatalytic events in this pathway. To identify these events, a soluble form of MT1-MMP, lacking the C-terminal transmembrane and cytoplasmic domains, was expressed in Pichia pastoris. Following secretion, the latent zymogen and active enzyme were each purified from media by fast protein liquid chromatography. Trace amounts of active MT1-MMP induced activation of the zymogen and its self-proteolysis. This autocatalytic processing generated six main forms of MT1-MMP, each of which was subjected to the N-terminal microsequencing to identify the cleavage sites. Our data indicate that MT1-MMP functions as a self-convertase and is capable of cleaving its own prodomain at the furin cleavage motif RRKR downward arrow Y(112), thus autocatalytically generating the mature MT1-MMP enzyme with an N terminus starting at Tyr(112). The mature enzyme undergoes further autocatalysis to the two distinct intermediates (N terminus at Trp(119) and at Asn(130)) and, next, to the three inactive ectodomain forms (N terminus at Thr(222), at Gly(284), and at Thr(299)). These findings provide, for the first time, a structural basis for understanding the unconventional mechanisms of MT1-MMP activation and regulation. Finally, our data strongly imply that MT1-MMP is a likely substitute for the general proprotein convertase activity of furin-like proteinases, especially in furin-deficient cancer cells.
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PMID:Membrane type-1 matrix metalloproteinase functions as a proprotein self-convertase. Expression of the latent zymogen in Pichia pastoris, autolytic activation, and the peptide sequence of the cleavage forms. 1251 92

Because beta-amyloid precursor protein (APP) has the abilities both to interact with extracellular matrix and to inhibit gelatinase A activity, this molecule is assumed to play a regulatory role in the gelatinase A-catalyzed degradation of extracellular matrix. To determine a region of APP essential for the inhibitory activity, we prepared various derivatives of APP. Functional analyses of proteolytic fragments of soluble APP (sAPP) and glutathione S-transferase fusion proteins, which contain various COOH-terminal parts of sAPP, showed that a site containing residues 579-601 of APP(770) is essential for the inhibitory activity. Moreover, a synthetic decapeptide containing the ISYGNDALMP sequence corresponding to residues 586-595 of APP(770) had a gelatinase A inhibitory activity slightly higher than that of sAPP. Studies of deletion of the NH(2)- and COOH-terminal residues and alanine replacement of internal residues of the decapeptide further revealed that Tyr(588), Asp(591), and Leu(593) of APP mainly stabilize the interaction between gelatinase A and the inhibitor. We also found that the residues of Ile(586), Met(594), and Pro(595) modestly contribute to the inhibitory activity. The APP-derived decapeptide efficiently inhibited the activity of gelatinase A (IC(50) = 30 nm), whereas its inhibitory activity toward membrane type 1 matrix metalloproteinase was much weaker (IC(50) = 2 microm). The decapeptide had poor inhibitory activity toward gelatinase B, matrilysin, and stromelysin (IC(50) > 10 microm). The APP-derived inhibitor formed a complex with active gelatinase A but not with progelatinase A, and the complex formation was prevented completely by a hydroxamate-based synthetic inhibitor. Therefore, the decapeptide region of APP is likely an active site-directed inhibitor that has high selectivity toward gelatinase A.
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PMID:Identification of a region of beta-amyloid precursor protein essential for its gelatinase A inhibitory activity. 1258 36

The MMP-11 proteinase, also known as stromelysin-3, probably plays an important role in human cancer because MMP-11 is frequently overexpressed in human tumors and MMP-11 levels affect tumorogenesis in mice. Unlike other MMPs, however, human MMP-11 does not cleave extracellular matrix proteins, such as collagen, laminin, fibronectin, and elastin. To help identify physiologic MMP-11 substrates, a phage display library was used to find peptide substrates for MMP-11. One class of peptides containing 26 members had the consensus sequence A(A/Q)(N/A) downward arrow (L/Y)(T/V/M/R)(R/K), where downward arrow denotes the cleavage site. This consensus sequence was similar to that for other MMPs, which also cleave peptides containing Ala in position 3, Ala in position 1, and Leu/Tyr in position 1', but differed from most other MMP substrates in that proline was rarely found in position 3 and Asn was frequently found in position 1. A second class of peptides containing four members had the consensus sequence G(G/A)E downward arrow LR. Although other MMPs also cleave peptides with these residues, other MMPs prefer proline at position 3 in this sequence. In vitro assays with MMP-11 and representative peptides from both classes yielded modest kcat/Km values relative to values found for other MMPs with their preferred peptide substrates. These reactions also showed that peptides with proline in position 3 were poor substrates for MMP-11. A structural basis for the lower kcat/Km values of human MMP-11, relative to other MMPs, and poor cleavage of position 3 proline substrates by MMP-11 is provided. Taken together, these findings explain why MMP-11 does not cleave most other MMP substrates and predict that MMP-11 has unique substrates that may contribute to human cancer.
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PMID:Identification of peptide substrates for human MMP-11 (stromelysin-3) using phage display. 1273 79

Human matrix metalloproteinase-26 (MMP-26/endometase/matrilysin-2) is a newly identified MMP and its structure has not been reported. The enzyme active site S1' pocket in MMPs is a well defined substrate P1' amino acid residue-binding site with variable depth. To explore MMP-26 active site structure-activity, a series of new potent mercaptosulfide MMP inhibitors (MMPIs) with Leu or homophenylalanine (Homophe) side chains at the P1' site were selected. The Homephe side chain is designed to probe deep S1' pocket MMPs. These inhibitors were tested against MMP-26 and several MMPs with known x-ray crystal structures to distinguish shallow, intermediate, and deep S1' pocket characteristics. MMP-26 has an inhibition profile most similar to those of MMPs with intermediate S1' pockets. Investigations with hydroxamate MMPIs, including those designed for deep pocket MMPs, also indicated the presence of an intermediate pocket. Protein sequence analysis and homology modeling further verified that MMP-26 has an intermediate S1' pocket formed by Leu-204, His-208, and Tyr-230. Moreover, residue 233 may influence the depth of an MMP S1' pocket. The residue at the equivalent position of MMP-26 residue 233 is hydrophilic in intermediate-pocket MMPs (e.g. MMP-2, -8, and -9) and hydrophobic in deep-pocket MMPs (e.g. MMP-3, -12, and -14). MMP-26 contains a His-233 that renders the S1' pocket to an intermediate size. This study suggests that MMPIs, protein sequence analyses, and molecular modeling are useful tools to understand structure-activity relationships and provides new insight for rational inhibitor design that may distinguish MMPs with deep versus intermediate S1' pockets.
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PMID:The intermediate S1' pocket of the endometase/matrilysin-2 active site revealed by enzyme inhibition kinetic studies, protein sequence analyses, and homology modeling. 1453 75

Leukolysin/membrane-type 6 matrix metalloproteinase (leukolysin/MT6-MMP), a glycosylphosphatidylinositol-anchored neutrophil matrix metalloproteinase, is also abnormally expressed in brain cancer tissues. Yet, little is known about its role in cancer progression. Here we show that MT6-MMP is capable of activating proMMP-2, an enzyme implicated in tumor invasion and metastasis. Although MT6-MMP is only 10% as active as MT5-MMP in mediating proMMP-2 activation, it generates a higher ratio of mature/intermediate forms of MMP-2 than MT5-MMP. Consistently, purified CAT of MT6-MMP converts proMMP-2 into mostly the mature form. Using the catalytically inactive mutant MMP-2EA (the E404A mutant of proMMP-2), which cannot autocatalytically mature from the intermediate form into the mature one, we show that MT6-MMP cleaves not only the known MT-MMP-processing site at Asn(66)-Leu but also the previously unsuspected Asn(109)-Tyr to yield a fully mature molecule. Despite their difference in mediating proMMP-2 activation in transfected cells, the CAT of MT6-MMP appears to be as efficient as that of MT5-MMP in cleaving proMMP-2EA in buffer, suggesting that its CAT is a strong proMMP-2 activator. Indeed, the CAT of MT6-MMP can partially substitute the CAT of prototypical MT1-MMP in mediating proMMP-2 activation. Taken these facts together, we conclude that MT6-MMP may participate in tumor invasion and metastasis by directly converting proMMP-2 into active form.
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PMID:Direct activation of pro-matrix metalloproteinase-2 by leukolysin/membrane-type 6 matrix metalloproteinase/matrix metalloproteinase 25 at the asn(109)-Tyr bond. 1458 71

Membrane-type 1 matrix metalloproteinase (MT1-MMP) and vascular endothelial growth factor (VEGF) are two key molecules involved in pericellular proteolysis and cell proliferation during tumor growth and angiogenesis. Our previous data showed that MT1-MMP overexpression in human breast carcinoma MCF7 cells induced an up-regulation of VEGF expression. This effect was associated in vivo with accelerated tumor growth and angiogenesis. We now provide evidence that MT1-MMP overexpression specifically affected VEGF-A production and failed to influence that of other VEGF family members (VEGF, B, C, D, or PlGF) or their receptors. The up-regulation of VEGF-A by MT1-MMP was related to an increased transcriptional activation rather than to a modification of mRNA stability. It was blocked by synthetic MMP inhibitors, TIMP2, but not TIMP-1 and abolished by a partial deletion of the catalytic domain or the cytoplasmic tail of MT1-MMP. Analysis of the signal transduction mechanisms demonstrated that MT1-MMP acts through a signaling pathway involving Src tyrosine kinases. Thus, our results provide new insight into the mechanisms of action of MT1-MMP during angiogenesis and suggest that the full enzymatic activity of MT1-MMP is required for a specific up-regulation of VEGF-A through an activation of Src tyrosine kinase pathways.
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PMID:Up-regulation of vascular endothelial growth factor-A by active membrane-type 1 matrix metalloproteinase through activation of Src-tyrosine kinases. 1472 79

The mechanisms by which c-erbB-dependent signaling contribute to the invasive potential of HNSCC remain to be fully elucidated. We have previously shown that c-erbB autocrine and/or paracrine stimulation upregulates MMP-9 but has no effect on the related gelatinase, MMP-2. BTC, a major c-erbB ligand, has the ability to efficiently activate all c-erbB receptors and to bind directly to EGFR and c-erbB-4. BTC is commonly expressed in HNSCC cells and exerts the most potent effects in terms of MMP induction relative to other c-erbB ligands so far tested. In the present study, we explored the contribution of major downstream events triggered by BTC/c-erbB receptor signaling to the regulation of MMP-9 and in vitro invasiveness of HNSCC cells. In human HNSCC cell lines, SIHN-006 and Detroit-562, BTC treatment resulted in rapid tyrosine phosphorylation of all c-erbB receptors whereas both endogenous MMP-9 and BTC-stimulated MMP-9 were predominantly mediated via EGFR. BTC induced ERK1/2, JNK/SAPK and Akt phosphorylation with differing kinetics but not p38 kinase. The BTC-dependent activation of JNK and PI3K/Akt pathways occurred predominantly via EGFR, whereas activation of the MEK-1/ERK pathway occurred via all 4 c-erbB receptors, although again predominantly via EGFR. Selective inhibition of ERK/MAPK (by PD98059 or U0126) and PI3K (by LY294002 or wortmannin) led to marked reduction of both basal and BTC-induced MMP-9 activity and invasive ability of HNSCC cells. In contrast, inhibition of p38 kinase with SB203580 produced no such effects. A specific inhibitor of NF-kappa B, BAY 11-7085, also blocked the stimulatory effect of BTC. No remarkable inhibition of MMP-9 and invasion was observed on targeting other cellular activities, such as PKA, PKC and PLC-gamma. Taken together, our data show that BTC induces MMP-9 production and invasion primarily through activation of EGFR, MAPK and PI3K/Akt in HNSCC cells.
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PMID:Signaling pathways required for matrix metalloproteinase-9 induction by betacellulin in head-and-neck squamous carcinoma cells. 1519 68

We have recently shown that stimulation of endothelial cells with vascular endothelial growth factor (VEGF) induces dissociation of caveolin-1 from the VEGFR-2 receptor, followed by Src family kinase-dependent tyrosine phosphorylation of the protein (Labrecque, L., Royal, I., Surprenant, D. S., Patterson, C., Gingras, D., and Beliveau, R. (2003) Mol. Biol. Cell 14, 334-347). In this study, we provide evidence that the VEGF-dependent tyrosine phosphorylation of caveolin-1 induces interaction of the protein with the membrane-type 1 matrix metalloproteinase (MT1-MMP). This interaction requires the phosphorylation of caveolin-1 on tyrosine 14 by members of the Src family of protein kinases, such as Src and Fyn, because it is completely abolished by expression of a catalytically inactive Src mutant or by site-directed mutagenesis of tyrosine 14 of caveolin-1. Most interestingly, the association of MT1-MMP with phosphorylated caveolin-1 induced the recruitment of Src and a concomitant inhibition of the kinase activity of the enzyme, suggesting that this complex may be involved in the negative regulation of Src activity. The association of MT1-MMP with phosphorylated caveolin-1 occurs in caveolae membranes and involves the cytoplasmic domain of MT1-MMP because it was markedly reduced by mutation of Cys574 and Val582 residues of the cytoplasmic tail of the enzyme. Most interestingly, the reduction of the interaction between MT1-MMP and caveolin-1 by using these mutants also decreases MT1-MMP-dependent cell locomotion. Overall these results indicate that MT1-MMP associates with tyrosine-phosphorylated caveolin-1 and that this complex may play an important role in MT1-MMP regulation and function.
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PMID:Src-mediated tyrosine phosphorylation of caveolin-1 induces its association with membrane type 1 matrix metalloproteinase. 1546 65

Matrilysin activity exhibits a broad bell-shaped pH-dependence profile, with pK(a) values of 4.0 and 9.8. A maximum of five out of eight tyrosine residues in matrilysin were nitrated with tetranitromethane. On nitration of between one and five tyrosines, pK(a) at the alkaline side (pK(e2)) was shifted from 9.8 to 10.3-10.6, while that at the acidic side (pK(e1)) was not altered. The pK(e2) that was shifted by nitration to 10.3-10.6 was restored to 9.4-9.7 by subsequent amination, suggesting that the shift in pK(e2) is induced by a negative charge introduced on the most reactive tyrosine, Tyr-150. The Michaelis constant (K(m)) observed at pH 10 was decreased by nitration as a result of the increase in pK(e2), suggesting that the residue with pK(e2) may play a role in the recognition of substrate. When four or five tyrosines were nitrated, the activity at pH <7 decreased significantly, while that at pH 7-10 was unchanged, and thus the pH-dependence was not bell-shaped, but anomalous, with a third pK(a) (pK(e3)) of 6.2-6.4 in addition to pK(e1) and pK(e2). This suggests the possibility that a newly introduced nitrotyrosine residue has a strong influence on the activity as an ionizable group.
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PMID:Anomalous pH-dependence of the activity of human matrilysin (matrix metalloproteinase-7) as revealed by nitration and amination of its tyrosine residues. 1548 74


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