2.8. Matrix metalloproteinases (MMPs)
MMPs comprise a family of at least 28 secreted or transmembrane enzymes collectively capable of processing and degrading various ECM proteins. Of these, at least 22 MMPs have so far been found to be expressed in human tissues. MMPs share high protein sequence homology and have defined domain structures and thus, according to their structural properties, MMPs are classified either as secreted MMPs or membrane anchored MMPs, which are further divided into eight discrete subgroups. Secreted MMPs include minimal-domain MMPs, simple hemopexin domain-containing MMPs, gelatin-binding MMPs, furin-activated secreted MMPs and vitronectin-like insert MMPs, while membrane bound MMPs include type I transmembrane MMPs, glycosyl-phosphatidyl inositol (GPI)-linked MMPs and type II transmembrane MMPs (see Fig. 2). (Reviewed in Egeblad & Werb 2002.)
Crystal structures of MMPs further uncovered the exact domain organization, polypeptide fold and main specificity determinants (reviewed in Bode et al. 1999). To date, crystal structures of the catalytic domains of human MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-11, MMP-12, MMP-13 and MMP-14, in addition to porcine full length MMP-1 and human proMMP-2 have been resolved (Van Doren et al. 1993, Reinemer et al. 1994, Bode et al. 1994, Lovejoy et al. 1994, Li et al. 1995, Fernandez-Catalan et al. 1998, Kiyama et al. 1999, Morgunova et al. 1999, Moy et al. 2000, Gall et al. 2001, Lang et al. 2001).
All MMPs are synthesized with a predomain containing a leader sequence, which targets the protein for secretion (reviewed in Sternlicht & Werb 2001). They are secreted as latent proforms, with a few exceptions of furin-processed proteinases, such as MMP-11 or MMP-28. The prodomain of MMPs has an egglike shape, and contains a well conserved cysteine switch motif of PRCXXPD for maintaining the proMMP latent (Springman et al. 1990, Van Wart & Birkedal-Hansen 1990). Generally, the structures of all MMP catalytic domains are quite similar (Bode et al. 1999). The shape of the catalytic domain is spherical with a flat active site cleft, which extends horizontally across the domain to bind peptide substrates or inhibitors. The catalytic domain has the zinc-binding motif, HEF/LGHS/ALGLXHS, which coordinates a zinc atom at the active site, and under the zinc, a ALMYP methionine-turn (Stöcker et al. 1995). The latency of the zymogen is maintained through cysteine-switch motif, in which the cysteine residue acts as a fourth zinc-binding ligand to maintain the enzyme inactive. In addition to the catalytic zinc, the catalytic domain also contains a structural zinc and two to three calcium ions. A sub-site- or S1’-pocket- or channel-like structure is a binding site for a substrate or inhibitor molecule within the active site, and differs considerably in size and shape among the various MMPs. P1’ indicates the residue of a bound substrate molecule. The P1’-S1’ interaction mainly determines the affinity of inhibitors, and the cleavage positions of peptide substrates. C-terminal hemopexin or vitronectin-like domains affect substrate or inhibitor binding, membrane activation and some proteolytic activities. The hemopexin domain, very similar in structure among the MMPs, is an ellipsoidal disc, and is connected to the catalytic domain by a hinge region. The hinge region is flexible and rich in proline residues. It may also influence substrate specificity. (Reviewed in Bode et al. 1999, Sternlicht & Werb 2001.)
Structural specificities among the different MMP subgroups can be recognized. Of the simple hemopexin domain-containing MMPs, which have the general structure of a preprodomain and a catalytic domain, which is connected through hinge to the hemopexin like-domain, at least MMP-1, MMP-8 and MMP-13 contain three conserved amino acids, Tyr-214, Asp-235, Gly-237, which are not present in other MMPs (Freije et al. 1994). Gelatin-binding MMPs have a unique 19 kDa fibronectin-like insert in the catalytic domain. This central domain is organized into three internal repeats, which are homologous to the type II motif of the collagen binding domain of fibronectin, and are needed for binding and cleaving collagen and elastin (Collier et al. 1988, 1992, Murphy et al. 1994, Shipley et al. 1996). MMP-9 has an additional, unique 54 amino acid long proline rich domain, not existing in other MMPs, which is homologous to the α2 chain of type V collagen (Wilhelm et al. 1989). Furin-activated secreted MMPs (MMP-11 and MMP-28) have a recognition motif for furin-like serine proteinases within their catalytic domain for intracellular activation. This motif is also found in the vitronectin-like insert MMPs (MMP-21), and the MT-MMPs (Egeblad & Werb 2002).
Type I transmembrane MMPs include MMP-14, MMP-15, MMP-16 and MMP-24, and have a carboxy-terminal, single-span transmembrane domain and a short cytoplasmic C-terminal tail (Kojima et al. 2000). MMP-17 and MMP-25 are anchored to the membrane by a C-terminal hydrophobic region with a glycosyl-phosphatidyl inositol (GPI) domain, and thus are classified as glycosyl-phosphatidyl inositol (GPI)-linked MMPs (Itoh et al. 1999, Kojima et al. 2000).
Figure 2. Domain structure of MMPs and their classification. MMPs are illustrated as latent zymogens with a catalytic site zinc (Zn) binding to cysteine (C) of cysteine-switch of the prodomain. (Modified from Egeblad & Werb 2002)
Both classes of carboxy-terminal domains localize all of these MT-MMPs in a very specific way to the cell surface, therefore playing an important role in diverse proteolytic events. MT-MMPs also have an eight amino acid insertion in the catalytic domain, which distinguish them from other MMPs (English et al. 2001). Analysis of this insertion in MMP-14 (163-PYAYIREG-170) indicates its importance for proMMP-2 activation (English et al. 2001).
Minimal-domain MMPs, MMP-7 and MMP-26, lack a hemopexin domain (Muller et al. 1988, de Coignac et al. 2000), whereas MMP-23 has cysteine array and immunoglobulin (Ig)-like domains instead of the conserved hemopexin-like domain (Pei et al. 2000). MMP-23 is also classified as type II transmembrane MMP, since it has an amino-terminal signal anchor (CA) targeting it to the cell membrane (Pei et al. 2000).
2.8.1. Simple hemopexin domain-containing MMPs
2.8.1.1. Collagenases
The story of MMPs begun when Gross and Lapiére (1962) detected collagenolytic activity in tissues of tadpoles, and called the respective enzyme collagenase. In oral tissues, this first characterized collagenase, MMP-1 (collagenase-1, interstitial collagenase, fibroblast collagenase), is detected in gingival fibroblasts capable of disrupting ECM collagen (Wilhelm et al. 1984). The MMP-1 gene has three dimorphic sites in the 5’ upstream region, which may affect the regulation of gene expression (Thiry-Blaise et al. 1995). The promoter region of the human MMP-1 gene contains AP-1 and ETS elements, which are able to mediate at least PMA induction (White et al. 1997). Analysis of the rabbit MMP-1 TGF-β inhibitory element (TIE) in the promoter region, which is also conserved in the human MMP-1 gene, reveals that the TIE represses constitutive MMP-1 gene transcription (White et al. 2000). The MMP-1 gene is transcribed into 2.5 kb mRNA (Wilhelm et al. 1986).
Latent MMP-1 has two major species of molecular mass, 57 kDa and 52 kDa, in a ratio of 1:4 in tissues (Wilhelm et al. 1984). The minor 57 kDa proMMP-1 is a result of N-glycosylations of the major 52 kDa form (Wilhelm et al. 1986). Active MMP-1 exists in forms of 48 kDa and 42 kDa, of which the 42 kDa MMP-1 represents the stable, active enzyme (Wilhelm et al. 1984, Grant et al. 1987). MMP-1 collagenolytic activity is essentially mediated through the (183)RWTNNFREY(191) motif of the catalytic domain in concert with the C-terminal hemopexin domain (Chung et al. 2000). Active MMP-1 hydrolyzes type I collagen into N-terminal , and the C-terminal fragments. However, it hydrolyzes type III collagen 10-fold faster than type I collagen (Wilhelm et al. 1984). MMP-1 also cleaves α-chains of native type II collagen (Fields et al. 1987), type VII collagen (Seltzer et al. 1989), type X collagen (Schmid et al. 1986), type VIII (Gadher et al. 1989), α2-macroglubulin (Sottrup-Jensen & Birkedal-Hansen 1989), gelatin and casein (Fields et al. 1990), α1-proteinase inhibitor and α1-antichymotrypsin (Desrochers et al. 1991), tenascin (Imai et al. 1994) and IL-1β (Ito et al. 1996).
MMP-8 (collagenase-2) represents the second collagenase, and was originally thought to be synthesised and stored exclusively in intracellular granules of human polymorphonuclear neutrophils (PMNs) in bone marrow (Bainton et al. 1971, Mainardi et al. 1991). MMP-8 has been purified from these granules, from which the PMNs secrete the enzyme, and it has been described as neutrophil type or polymorphonuclear type (PMN) MMP-8 (Hasty et al. 1986). Mesenchymal type MMP-8, differing from neutrophil MMP-8 in protein size, is expressed by human chondrocytes (Cole et al. 1996), rheumatoid synovial fibroblasts and endothelial cells (Hanemaaijer et al. 1997). It is also produced in keratinocytes, including oral squamous cell carcinoma cells (Bachmeier et al. 2000, Moilanen et al. 2002) and plasma cells (Wahlgren et al. 2001). Furthermore, MMP-8 has been found in human gingiva (Tonetti et al. 1993), saliva (Ingman et al. 1994), dental plaque (Sorsa et al. 1995) and demineralised dentinal caries lesions (Tjäderhane et al. 1998a).
A 3.3 kb MMP-8 mRNA species codes for both neutrophil and mesenchymal type protein forms (Hasty et al. 1990, Hanemaaijer et al. 1997). It has been reported that a splice variant of the MMP-8 transcript exists. A cDNA with 91 bp insertion in its sequence still codes for the active MMP-8 protein. However, it is not secreted from cells (Hu et al. 1999). MMP-8 shares 57% identity to the MMP-1 protein sequence (Hasty et al. 1990). It contains eight possible glycosylation sites (Hasty et al. 1990), and hence several molecular weight forms of the MMP-8 protein have been published. The PMN type proMMP-8 has been reported to be 75–85 kDa in size. Active MMP-8 may be of 45 kDa, 57 or 64 kDa, respectively, in addition to smaller 22–28 kDa species (Hasty et al. 1986, Tschesche et al. 1992). The size of the chondrocyte proMMP-8 is 55 kDa and active forms are 46 and 42 kDa (Cole et al. 1996). MMP-8, secreted from the rheumatoid synovial fibroblasts and endothelial cells, is 50 kDa in size (Hanemaaijer et al. 1997).
MMP-8 degrades type I collagen into characteristic and fragments (Hasty et al. 1987, 1990). It also cleaves types II and III collagen molecules, yet with slower catalytic rates than type I collagen (Hasty et al. 1987). MMP-8 may also cleave ECM molecules other than triple helix collagens. It is able to cleave aggrecan (Arner et al. 1997), plasma serine-proteinase inhibitor, α1-antitrypsin (Michaelis et al. 1990) and fibrinogen (Hiller et al. 2000).
MMP-13 (collagenase-3) was originally cloned from human breast tumour (Freije et al. 1994). The expression of MMP-13 is not restricted to breast tumours, but is also expressed by human chondrocytes (Blavier & Delaisse 1995, Mitchell et al. 1996, Reboul et al. 1996, Borden et al. 1996), synovial membrane (Wernicke et al. 1996), synovial stroma (Lindy et al. 1997), synovial fibroblasts (Westhoff et al. 1999), gingival fibroblasts (Ravanti et al. 1999) and plasma cells (Wahlgren et al. 2001). It is also expressed by hypertrophic chondrocytes, osteoblasts, periosteal cells and fibroblasts during human fetal bone development (Johansson et al. 1997, Stahle-Backdahl et al. 1997), and postnatally in bone remodeling (Stahle-Backdahl et al. 1997).
MMP-13 is transcribed into three different mRNA species with respective sizes of 2, 2.5 and 3 kb (Freije et al. 1994, Reboul et al. 1996). The human MMP-13 gene promoter contains recognition sites for TATA and CCAAT DNA-binding proteins, an AP-1 motif, PEA-3 consensus sequence, an osteoblast specific element (OSE-2), a TGF-β inhibitory element (TIE) and three motifs of hormone response elements (Pendas et al. 1997a, Tardif et al. 1997). Co-operation of SMAD proteins with the functional AP-1 site, in concert with PEA-3, is essential for both basal and inducible gene transcription, e.g. by TGF-β in chondrocytes (Tardif et al. 2001). MMP-13 has an AG-rich element (AGRE) in the proximal promoter region, which represses basal transcription of the gene (Benderdour et al. 2002). A cDNA of MMP-13 codes for a 471 amino acid polypeptide. It has three potential N-glycosylation sites within an active protein molecule (Freije et al. 1994). The molecular weight of the proMMP-13 is 60–65 kDa. Active MMP-13 is 50–55 kDa in size (Freije et al. 1994, Knäuper et al. 1996a), but is further cleaved into a final active form of 48 kDa (Knäuper et al. 1996a).
Active human MMP-13 preferentially hydrolyses fibrillar type II collagen (Knäuper et al. 1996a, Mitchell et al. 1996, Reboul et al. 1996), and type II procollagen, both at the telopeptide and N-proteinase sites (Fukui et al. 2002). It also cleaves gelatin (Knäuper et al. 1996a) and cleaves type I and type III collagens into characteristic N-terminal and C-terminal fragments (Freije et al. 1994, Knäuper et al. 1996a). MMP-13 cleaves type II collagen approximately 10 times faster (Mitchell et al. 1996) and gelatin about 40 times more efficiently than MMP-1 or MMP-8, while type I collagen is cleaved with comparable efficiency to the MMP-1 and MMP-8 (Knäuper et al. 1996a). In addition, MMP-13 degrades types IV, IX, X and XIV collagens, tenascin, fibronectin, fibronectin fragments (Knäuper et al. 1997), cartilage proteoglycan, aggrecan (Fosang et al. 1996), plasma proteins, fibrinogen and Factor XII (Hiller et al. 2000).
MMP-18 (collagenase-4) was identified in Xenopus laevis (Stolow et al. 1996), but a human counterpart for this amphibian collagenase has not yet been identified. Xenopus collagenase-4 is 54% identical with human MMP-1. It contains the highly conserved cysteine-switch motif, except that proline is replaced with tyrosine. It degrades gelatine, and cleaves type I collagen into characteristic and fragments. (Stolow et al. 1996.)
2.8.1.2. Stromelysins
MMP-3 (stromelysin-1) was purified and characterized from human rheumatoid synovial fibroblasts (Okada et al. 1986) and skin fibroblasts (Wilhelm et al. 1987). The promoter of MMP-3 contains three elements important for mitogenic induction: a SPRE site (stromelysin-1 PDGF-responsive element) (Sanz et al. 1994), a PEA3 site (polyomavirus enhancer A-binding protein-3 site) (Wasylyk et al. 1991, Buttice & Kurkinen 1993) and an AP-1 site (activator protein-1 binding site) (Kerr et al. 1988). SPRE is the binding site for a transcription factor, SPBP (SPRE-binding protein), which has the ability to enhance transcription of other transcription factors such as c-Jun, Ets1, Sp1 and Pax6 (Rekdal et al. 2000). The PEA-3 site mediates TPA induction of human MMP-3 gene transcription (Buttice & Kurkinen 1993). The AP-1 site in the promoter of MMP-3 mediates basal gene expression, but is not necessary for the PMA-response of human MMP-3 (Buttice et al. 1991). Animal studies have revealed that the TGF-β 1 inhibitory element (TIE) in the promoter region mediates the TGF-β 1 inhibitory effect on stromelysin gene expression (Kerr et al. 1990). However, the age of the cell population affects whether MMP-3 is induced or repressed by TGF-β 1. In young fibroblasts, TGF-β 1 represses the gene induction, whereas in older cells, repression is not apparent (Edwards et al. 1996).
MMP-3 is 54% identical to human MMP-1 and 87% homologous to its rat counterpart (Saus et al. 1988). ProMMP-3 is secreted as a 57 kDa form, which can be glycosylated into a 60 kDa protein (Wilhelm et al. 1987). Latent MMP-3 is processed into a 53 kDa transient intermediate form by cleaving 35 amino acids of the propeptide. Autolysis of the intermediate form yields the 45 kDa mature active MMP-3, which may further be processed to a smaller active form of 28 kDa (Okada et al. 1986, Freimark et al. 1994). Latent and active high molecular forms of MMP-3 can bind to collagen fibrils, and to other ECM components via their C-terminal domains, whereas active low molecular forms do not (Allan et al. 1991). MMP-3 in synovia of osteo- and rheumatoid arthtritis is bound either to ECM or is located in cells (Hembry et al. 1995). MMP-3 is also bound into the fibrous tissue and osteoid of human bone tissue (Bord et al. 1999).
Active MMP-3 is at least capable of degrading cartilage proteoglycans, gelatin, type IV collagen, laminin, fibronectin (Okada et al. 1986) and type IX collagen (Okada et al. 1989). However, MMP-3 digests fibronectin and gelatin more efficiently at an acidic pH (pH 5.5) (Gunja-Smith et al. 1989). MMP-3 is able to remove the NH2-terminal propeptide from type I procollagen, and in the case of heat denatured type I procollagen also the COOH-terminal propeptide (Okada et al. 1986). It also cleaves type III procollagen (Murphy et al. 1991). Recombinant human stromelysin-1 acts as a telopeptidase against types II and XI collagens, and hydrolyses collagen type X (Wu et al. 1991) and aggrecan (Flannery et al. 1992). Furthermore, MMP-3 degrades fibrinogen and fibrin (Bini et al. 1996), IL-1β (Ito et al. 1996), decorin (Imai et al. 1997), urokinase-type plasminogen activator (Ugwu et al. 1998) and plasminogen activator inhibitor-1 (PAI-1) (Lijnen et al. 2000). ProMMP-3 may contribute to plasminogen activation through complexing with both plasminogen and tissue-type plasminogen activator (t-PA), rather than hydrolysing t-PA (Arza et al. 2000). MMP-3 also cleaves serpin α2-antiplasmin (Lijnen et al. 2001) and IgG (Gearing et al. 2002) and type II procollagen both at the telopeptide and N-proteinase sites (Fukui et al. 2002).
The second human stromelysin, MMP-10 (stromelysin-2), was cloned from rheumatoid synovial fibroblasts (Sirum & Brinckerhoff 1989). Other human cells such as keratinocytes (Windsor et al. 1993), T-lymphocytes (Conca & Willmroth 1994), chondrocytes, osteoblasts and osteoclasts (Bord et al. 1999) also produce MMP-10. The MMP-10 gene contains elements for PMA, EGF and IL-1β induction within its promoter region (Sirum & Brinckerhoff 1989). MMP-10 is transcribed into 1.8 kb mRNA (Conca & Willmroth 1994). Latent MMP-10 is 54 kDa, and after proteolytic activation has a molecular mass of 44 kDa (Windsor et al. 1993). MMP-10 is 82% similar to MMP-3 (Sirum et al. 1989). Active MMP-10 cleaves collagen types III, IV and V, fibronectin and gelatin (Nicholson et al. 1989). It also cleaves proteoglycan in both acid and neutral environments (Fosang et al. 1991).
2.8.1.3. Others
Human MMP-12, also called macrophage metalloelastase, is mainly expressed by alveolar macrophages (Shapiro et al. 1993). Generally, it is not expressed at detectable levels in normal adult tissues. However, MMP-12 expression is induced as a response to inflammatory stimuli such as endotoxin, or in remodeling tissues like the placenta (Belaaouaj et al. 1995). The promoter sequence of MMP-12 contains a TATA-box, AP-1 motif and a PEA3 element (Belaaouaj et al. 1995). The 54 kDa proMMP-12 is processed to an active enzyme of 22 kDa, which degrades elastin (Shapiro et al. 1993). MMP-12 is also able to degrade type IV collagen, gelatin, fibronectin, laminin, vitronectin, proteoglycan and myelin basic protein. It cleaves α1-antitrypsin and releases tumour necrosis factor (TNF) from a proTNF fusion protein (Chandler et al. 1996). MMP-12 also cleaves tissue factor pathway inhibitor (Belaaouaj et al. 2000), fibrinogen and Factor XII (Hiller et al. 2000), and urokinase-type plasminogen activator receptor (uPAR) (Koolwijk et al. 2001).
MMP-19, cloned from human mammary gland, was at first designated as MMP-18 (Cossins et al. 1996). However, Xenopus collagenase-4 represents the 18th MMP. Furthermore, it was found that MMP isolated from a liver cDNA library (Pendas et al. 1997b), was identical to the mammary gland MMP, and was thus designated as the 19th MMP, MMP-19. MMP-19 has also been isolated from synovial blood vessels of a rheumatic arthritis patient with the name RASI-1 (Kolb et al. 1997). MMP-19 differs from other known MMPs in that it has an insertion of acidic amino acids at the hinge region, and two potential glycosylation sites in the hemopexin domain (Cossins et al. 1996, Pendas et al. 1997b). In addition, proline-94 is replaced with a glutamic acid in the conserved cysteine-switch motif (Cossins et al. 1996). MMP-19 is able to associate with the cell surface through its hemopexin domain (Mauch et al. 2002). In addition to several internal organs (Pendas et al. 1997b), MMP-19 is expressed at the surface of blood mononuclear cells, lymphocytes (Sedlacek et al. 1998), blood vessels (Kolb et al. 1997, 1999), normal breast tissue, and both malignant and benign breast lesions (Djonov et al. 2001), lung fibroblasts and at the myeloid cell surface (Mauch et al. 2002).
The MMP19 promoter has several cis-acting regulatory elements similar to other MMPs, including a TATA-box, an AP-1 binding site and a binding site for transcription factors of the Ets family (PEA3 element) (Mueller et al. 2000). MMP-19 full lenght cDNA encodes a 508 residue polypeptide with predicted molecular weight of 57 kDa (Cossins et al. 1996, Pendas et al. 1997b). MMP-19 degrades gelatin (Sedlacek et al. 1998), type IV collagen, laminin, nidogen, tenascin-C, fibronectin, type I gelatin (Stracke et al. 2000b), cartilage aggrecan and oligomeric matrix protein (COMP) (Stracke et al. 2000a).
In mature tissues, MMP-20 (enamelysin) is almost exclusively expressed by odontoblasts, cells capable of forming tooth dentin matrix, and to a lesser extent in pulp tissue (Llano et al. 1997, Sulkala et al. 2002). During tooth development MMP-20 localizes to ameloblasts, odontoblasts and enamel and to a lesser extent to dentin (Bartlett et al. 1996, Den Besten et al. 1998, Caterina et al. 1999, 2000, Takata et al. 2000). Recently, MMP-20 was found to be expressed in oral squamous cell carcinoma (SCC) cells in vitro (Väänänen et al. 2001), but to date they are the only cells outside the tooth-forming cells that have been shown to express MMP-20, despite various attempts. The MMP-20 transcript utilizes two alternative polyadenylation sites, giving two MMP-20 transcripts of sizes of 2 kb and 4 kb, respectively (Bartlett et al. 1996). The open reading frame of the MMP-20 cDNA codes for a 483 amino acid protein. Latent MMP-20 has a molecular weight of 54.4 kDa and the active form is 42.6 kDa (Bartlett et al. 1996, Llano et al. 1997, Li et al. 1999). In addition, 42.6 kDa active forms may be further cleaved into several smaller enzymes ranging from 18 kDa to 38 kDa, possessing differential catalytic activities (Den Besten et al. 1998, Ryu et al. 1999, Li et al. 1999). MMP-20 differs from other MMPs in that it has an insertion of basic residues in the hinge region. The protein sequence does not contain any glycosylation sites (Llano et al. 1997). MMP-20 cleaves amelogenin (Llano et al. 1997), gelatin, casein, aggrecan and cartilage oligomeric matrix protein (COMP) (Fukae et al. 1998, Ryu et al. 1999, Stracke et al. 2000a), fibronectin, type IV collagen, laminin-1 and -5 and tenascin-C (Väänänen et al. 2001).
MMP-22 (CMMP) has been cloned from chicken embryo fibroblasts (Yang & Kurkinen 1998). A human homologue has not been identified. MMP-22 is a 472 amino acid polypeptide with a unique cysteine in the catalytic domain, which is at the same position in the catalytic domains of both MMP-19 and XMMP. The calculated molecular weight of the active CMMP is 42 kDa. Recombinant MMP-22 digests both gelatin and casein with equal efficiency as recombinant MMP-1. (Yang & Kurkinen 1998.)
The MMP-27 1655 bp cDNA has been cloned by Benoit de Coignac et al. (unpublished), and submitted to the nucleotide genbank of NCBI with the accession number of AF195192. It has not yet been further characterized or published.
2.8.2. Gelatin-binding MMPs
This subgroup of metalloproteinases was originally described as type IV collagenases, because of their ability to cleave type IV collagen. One of these, MMP-2 (gelatinase-A, 72 kDa type IV collagenase) was originally purified from highly a metastatic murine tumour (Liotta et al. 1981, Salo et al. 1985). Since then, it has been found to be expressed in several normal and malignant human tissues. In oral tissues, gingival fibroblasts express MMP-2 (Hipps et al. 1991). In human hard tissues, MMP-2 is produced by osteoblasts and odontoblasts (Tjäderhane et al. 1998b, Rifas et al. 1989). MMP-2 has also been identified in sound (Martin-De Las Heras et al. 2000) and in carious (Tjäderhane et al. 1998a) human dentin.
The MMP-2 gene codes for a single mRNA of 3.1 kb (Collier et al. 1988). The promoter of the human MMP-2 gene has no TATA box or TPA/PMA responsive element (Huhtala et al. 1990). Nor does it have a CAAT box, but a binding site for the transcription factor AP-2 exists in the first exon (Huhtala et al. 1990). Further MMP-2 sequence analysis has revealed other important transcription factor binding sites in the promoter region, such as AP-1, PEA3, C/EBP, CREB, Ets-1 and Sp1 sites (Qin et al. 1999). In addition, the MMP-2 promoter has a binding site for the tumour suppressor and transcription factor, p53 (Bian & Sun 1997). The Sp1 and AP-2 elements have been reported to mediate constitutive human MMP-2 expression (Qin et al. 1999), whereas interaction of AP-2 with the transcription factor YB-1 within an enhancer element (RE-1) induces MMP-2 gene transcription (Mertens et al. 1998). Further characterization reveals two potential N-linked glycosylation sites at Asn546 and Asn613. However, no N-linked oligosaccaharides have been found with the MMP-2 protein (Collier et al. 1988). The molecular weight of proMMP-2 is 70 to 72 kDa, and active species are 65 kDa and 62 kDa (Stetler-Stevenson et al. 1989b, Hipps et al. 1991).
MMP-2 preferentially cleaves gelatin and type IV collagen, but also types V and VII collagens and fibronectin (Collier et al. 1988), and type X collagen (Welgus et al. 1990). MMP-2 is also able to cleave soluble, triple helical type I collagen at the typical Gly-Ile/Leu sites, producing the and fragments (Aimes & Quigley 1995). The catalytic and hemopexin domains of MMP-2, but not the fibronectin domain, are responsible for collagenolysis (Patterson et al. 2001). In addition, MMP-2 degrades cartilage proteoglycan and elastin (Okada et al. 1990), IL-1β (Ito et al. 1996) decorin (Imai et al. 1997) and laminin-5 (Giannelli et al. 1997).
MMP-2 exists in the ECM bound to type I and type IV collagen molecules, gelatin and laminin. MMP-2 binds to type I collagen through the fibronectin domain, which stabilises it from autolysis, thereby controlling its activity (Allan et al. 1995, Ellerbroek et al. 2001). On the other hand, latent MMP-2 is one of the few MMPs so far known to localize to the cell membrane for proteolytic activation (Sato et al. 1994).
The second gelatinase MMP-9 (92 kDa type IV collagenase, gelatinase B) is produced in human macrophages and polymorphonuclear leukocytes (Murphy et al. 1989). It has also been localized into the endothelial cells and synovial fibroblasts in rheumatoid arthritis synovium (Ahrens et al. 1996). MMP-9 is expressed by osteoclasts in the human normal bone tissues, implicating a role in the bone remodeling (Okada Y et al. 1995). Mature human intact odontoblasts also express MMP-9 (Tjäderhane et al. 1998b). In addition, it has been identified in human dental caries lesion (Tjäderhane et al. 1998a) and saliva (Davis 1991). However, MMP-9 is not expressed by human gingival fibroblasts (Bolcato-Bellemin et al. 2000). Like MMP-2, MMP-9 may exist in the ECM bound to type I collagen, gelatin or laminin. (Allan et al. 1995). Localization of the activated MMP-9 to the cell surface by the hyaluronan receptor CD44 may mediate the activation of latent TGF-β by MMP-9 (Bourguignon et al. 1998, Yu & Stamenkovic 2000).
The MMP-9 gene is transcribed into a 2.5 kb mRNA species (Huhtala et al. 1991). The 5’ flanking region of the gene contains binding sites for AP-1, NF-κ B, and Sp1, which synergistically mediate the induction of MMP-9 gene expression by TPA or TNF-α, and TGF-β inhibitor element (TIE) (Huhtala et al. 1991, Sato & Seiki 1993). The GT box located downstream of the AP-1 site is essential for the induction of gene transcription by v-Src, which is also able to mediate promoter activation via the AP-1 site (Sato et al. 1993). Ets and Sp-1 are essential for activation of MMP-9 gene expression in fibroblasts (Himelstein et al. 1998). NF-κ B is necessary for the upregulation of MMP-9 gene by inflammatory cytokines, IL-1α or TNF-a, but not by bFGF or PDGF (Bond et al. 1998). AP-1 slightly mediates the gene transcription by bFGF, PDGF, IL-1α or TNF-α (Bond et al. 1998). Functional polymorphism in the promoter of the MMP-9 gene results in variation in its expression at the transcriptional level (Peters et al. 1999).
MMP-9 is synthesized as a 78.4 kDa prepropeptide, and is secreted as a glycosylated 92 kDa proenzyme (Wilhelm et al. 1989). Proteolytic activation of the zymogen yields an active MMP-9 enzyme of 82 kDa (Ogata et al. 1992). MMP-9 may exist as a monomer, homodimer, or as a complex with lipocalin in neutrophils (Kolkenbrock et al. 1996). Active MMP-9 hydrolyses gelatin, native type IV collagen, elastin (Murphy et al. 1991), α2 chain of type I collagen, native collagens types III, V, XI and XIV (Okada et al. 1992, O’Farrell & Pourmotabbed 1998) and type II procollagen both at the telopeptide and N-proteinase sites (Fukui et al. 2002). MMP-9 is also able to cleave the cartilage proteoglycan, aggrecan, although very slowly compared to MMP-2, MMP-3 or MMP-7 (Fosang et al. 1992). MMP-9 hydrolyses human plasminogen generating angiostatin fragments (Patterson & Sang 1997), and degrades IL-1β (Ito et al. 1996). One unique feature of MMP-9 is that latent MMP-9 (purified from placenta tissue sections) is catalytically active against both the fluorogenic peptide MCA-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 and gelatin substrates (Bannikov et al. 2002).
2.8.3. Furin-activated secreted MMPs
MMP-11, or stromelysin-3, was originally characterized from breast carcinoma and has been shown to be 36–40% similar with MMP-3 or MMP-10 or MMP-1 (Basset et al. 1990). The promoter of the MMP-11 gene does not contain an AP1 element, which typically exists in this group of several other MMPs, but instead contains a DR1 retinoic acid responsive element (Anglard et al. 1995), and is induced by retinoic acid in human fibroblasts (Guerin et al. 1997). The MMP-11 promoter contains a TPA-responsive element, required for basal gene expression, and C/EBP-binding site for inducible gene regulation (Luo et al. 1999). MMP-11 expression is induced by TGF-β in osteoblasts and fibroblasts (Delany & Canalis 2001).
A recombinant MMP-11 is cleaved to a range of different sizes from 20 to 65 kDa (Pei et al. 1994). Of these, the 45 kDa species exhibits catalytic activity capable of cleaving α2-macroglobulin and α1-proteinase inhibitor, which inhibits serine proteinases (Pei et al. 1994). MMP-11 is also able to cleave insulin-like growth factor-binding protein-1 (IGFBP-1) indicating a possible role in regulating availability of IGF-1 (Manes et al. 1997). No other substrates have been found for MMP-11 to date.
The recently identified MMP-28 (epilysin) cDNA codes for a 520 amino acid protein with molecular mass of 59 kDa (Lohi et al. 2001, Marchenko & Strongin 2001). It is expressed in testis, lungs, heart, colon, intestine, brain and skin, fetal kidney, and in several tumour types (Lohi et al. 2001, Marchenko & Strongin 2001). The MMP-28 promoter contains Sp1/Sp3 binding GT-box, but does not contain the TATA-box and CAAT sequence typical for most MMPs (Illman et al. 2001). MMP-28, capable of degrading at least casein, shares 40% homologue with MMP-19. The cysteine-switch motif of the MMP-28 has a threonine-94 residue instead of proline-94, present in all other MMPs except MMP-19. MMP-28 also has an 11 amino acid insertion after cysteine-switch motif followed by an RRKKR furin recognition site. (Lohi et al. 2001, Marchenko & Strongin 2001.)
2.8.4. Vitronectin-like insert MMPs
MMP-21 has been partially cloned from human multi-tissue gene library (Marchenko et al. 2001). The 1038 bp fragment encodes a partial sequence of the putative catalytic domain followed by the putative hinge and hemopexin domain. This partial sequence of the human MMP-21 is 73% identical to the Xenopus XMMP/MMP-21. Xenopus XMMP has 604 amino acids including a putative 22-residue signal peptide (Yang et al. 1997). Characteristic of XMMP is that located in its propeptide domain is a 37 amino acid-long insertion domain (ID), related to vitronectin, that has not been found in other MMPs. In addition, it lacks a proline-rich hinge region between the catalytic and hemopexin/vitronectin-like domain (Yang et al. 1997).
2.8.5. Minimal-domain MMPs
MMP-7 is also known as PUMP-1 (putative metalloproteinase) or matrilysin-1. Human MMP-7 cDNA was cloned from tumours (Muller et al. 1988). It is expressed in osteoarthritic cartilage (Ohta et al. 1998), and is tightly bound into ECM heparin sulfate proteoglycan (Yu & Woessner 2000). The gene promoter of MMP-7 contains TATA, AP-1 and PEA3 elements (Gaire et al. 1994). Latent MMP-7 has molecular weight of 28 kDa, which can be activated by APMA into active species of 21 and 19 kDa (Quantin et al. 1989). MMP-7 has broad substrate spectrum, and it is capable of degrading several components, some of which also exist in dental tissue. These include fibronectin, gelatin, proteoglycan, decorin, tenascin-C, laminin, osteonectin, osteopontin, E-cadherin, proTNF-α and IgG (Quantin et al. 1989, Miyazaki et al. 1990, Murphy et al. 1991, Gearing et al. 1994, 2002, Siri et al. 1995, Imai et al. 1997, Sasaki et al. 1997, Agnihotri et al. 2001, Noe et al. 2001, Davies et al. 2001). MMP-7 also fragments corneal collagen type XVIII NC1 domain, generating 28 kDa endostatin (Lin HC et al. 2001), and type II procollagen (Fukui et al. 2002).
Minimal domain MMP-26 (matrilysin-2, endometase) was simultaneously cloned from a fetal cDNA library (de Coignac et al. 2000), human endometrial tumours (Park et al. 2000) and placenta (Uria & Lopez 2000). It has a threonine residue adjacent to the Zn-binding site characteristic of matrilysins (Park et al. 2000, Uria & Lopez 2000). MMP-26 shares 39% homology to MMP-7 (Park et al. 2000). The MMP-26 gene is located chromosome 11p5 (Uria & Lopez 2000) and is transcribed into an 1.03 kb size mRNA species (Park et al. 2000). Latent MMP-26 has a molecular weight of 28–29 kDa and active MMP-26 is 19 kDa (de Coignac et al. 2000, Park et al. 2000, Uria & Lopez 2000). MMP-26 cleaves gelatin, α1-proteinase inhibitor, TNF-α convering enzyme (Park et al. 2000), type IV collagen, fibronectin, fibrinogen (Uria & Lopez 2000), β -casein (de Coignac et al. 2000) and vitronectin (Marchenko et al. 2001).
ProMMP-26 contains a unique histidine residue instead of arginine residue in the cysteine-switch motif (PH(81)CGXXD) (Park et al. 2000, Marchenko et al. 2001, Marchenko et al. 2002). Since PH(81)CGXXD is not functional, proMMP-26 is not activated by conventional pathway, although His(81)Arg(81) mutation restores functionality to the cysteine-switch motif in the prodomain of MMP-26 (Marchenko et al. 2002). Marchenko and colleagues (2002) further show that autolytic LLQ(59)(60)QFH cleavage upstream of the cysteine-switch motif induces the proteolytic activity of recombinant proMMP-26, indicating an alternative activation pathway for this proenzyme.
2.8.6. Type I transmembrane MMPs
MMP-14 (MT1-MMP) was originally discovered as being able to activate proMMP-2 on the cell surface of invasive lung carcinoma cells (Sato et al. 1994). In mineralized tissues, MMP-14 is expressed by mouse osteoblasts (Mizutani et al. 2001), rabbit osteoclasts (Sato et al. 1997), and by odontoblasts and ameloblasts of developing porcine tooth tissues (Caron et al. 1998), and in vitro also by human osteoblast-like cells (Luo & Liao 2001). The MMP-14 gene promoter contains several regulatory elements, such as four CCAAT-boxes and one Sp-1 site, but no TATA-box (Lohi et al. 2000). MMP-14 mRNA is 4.5 kb and encodes a 582 amino acid protein (Sato et al. 1994). The molecular mass for latent MMP-14 is 63–66 kDa and for active MMP-14 is 54 kDa (Sato et al. 1994, Sang and Douglas, 1996). The active enzyme may further be processed into an inactive 43 kDa form (Lohi et al. 1996). MMP-14 molecules form a multimeric complex with each other through hemopexin and cytoplasmic domains, which mediates its autocatalytic processing and proMMP-2 activation (Itoh et al. 2001, Lehti et al. 2002). The propeptide domain of MMP-14 may act as an intramolecular chaperone for efficient trafficking of MT1-MMP to the cell surface (Cao et al. 2000, Pavlaki et al. 2002). MMP-14 has two potential proprotein convertase recognition motifs, RRPR/RRKR, for possible intracellular furin activation of the zymogen (Sato et al. 1994, Will & Hinzmann 1995, Yana & Weiss 2000).
Native and transmembrane deleted MMP-14 cleaves types I, II and III collagens (d’Ortho et al. 1997, Ohuchi et al. 1997). Soluble MMP-14 also cleaves gelatin, cartilage proteoglycan, fibronectin, vitronectin, tenascin, nidogen, aggrecan, perlecan, laminin-1, α1-proteinase inhibitor and α2-macroglobulin (Pei & Weiss 1996, d’Ortho et al. 1997, Ohuchi et al. 1997) and type II procollagen both at the telopeptide and N-proteinase sites (Fukui et al. 2002). MMP-14 is able to cleave fibrinogen and also inactivate Factor XII (Hiller et al. 2000). It degrades the hyaluronan receptor CD44 (Kajita et al. 2001), the receptor of complement component (gC1qR) (Rozanov et al. 2001) and the cell surface adhesion receptor, tissue transglutaminase (tTG) (Belkin et al. 2001). MT1-MMP is capable of processing pro-αV integrin, thus exhibiting integrin convertase activity (Deryugina et al. 2001, 2002a, Ratnikov et al. 2002). MMP-14 complexed to TIMP-2 possesses gelatinolytic activity (Imai et al. 1996). MMP-14 on the cell membrane is able to degrade type I collagen more efficiently after addition of proMMP-2 (Atkinson et al. 2001).
MMP-14 may itself act as a regulatory protein since the cytoplasmic domain of MMP-14 activates extracellular signal-regulated protein kinase (ERK), indicating a role in a signal transduction pathway involved in cell migration or gene regulation (Gingras et al. 2001). MMP-14 up-regulates vascular endothelial growth factor (VEGF) in human glioma U251 xenografts in athymic mice (Deryugina et al. 2002b).
The MMP-15 (MT2-MMP) 3.6 kb transcript was originally characterised in human lung and shown to be expressed by several other internal organs (Will & Hinzmann 1995). MMP-15 is 75.8 kDa in size, containing a possible glycosylation site at N150. It is 73.9% similar with MMP-14 (Will & Hinzmann 1995). Recombinant catalytic domain of MMP-15 cleaves fibronectin, tenascin, nidogen, aggrecan, perlecan and laminin (d’Ortho et al. 1997). MMP-15 degrades the cell surface adhesion receptor, tissue transglutaminase (tTG) (Belkin et al. 2001).
MMP-16 (MT3-MMP) was cloned from a placenta cDNA library (Takino et al. 1995). It is also expressed in the brain at high levels. The MMP-16 transcript is 12 kb in size, encoding a polypeptide of 604 amino acids (Takino et al. 1995). ProMMP-16 molecular mass is 64 kDa. Smaller, possibly active forms of 52, 33 and 30 kDa have also been characterised. Soluble MMP-16 is obtained by an alternative mRNA splicing (Matsumoto et al. 1997). MMP-16 also has a RXKR motif for furin processing. Its catalytic domain shares 66% homology with MMP-14 (Takino et al. 1995), and it is also capable of activating proMMP-2 (Takino et al. 1995). Soluble MMP-16 hydrolyses type III collagen, α2(I) collagen chain, cartilage proteoglycan, gelatin, fibronectin, vitronectin, laminin-1, transferrin, α1-proteinase inhibitor and α2-macroglobulin (Matsumoto et al. 1997, Shimada et al. 1999). MMP-16 also degrades the cell surface adhesion receptor, tissue transglutaminase (tTG) (Belkin et al. 2001).
Human MMP-24 (MT5-MMP), encoding a 645 amino acid polypeptide, is expressed in the brain, kidney, pancreas and lung. The cleavage site for shedding of MMP-24 is located at (545) RRKERR, where a furin recognition site also exists, and therefore proteolytic activity of MMP-24 could be regulated by shedding with furin-type convertase (Wang & Pei 2001). However, the substrates for MMP-24 remain largely unknown.
2.8.7. GPI-linked MMPs
MMP-17 (MT4-MMP) is widely expressed in the internal organs and by leukocytes (Puente et al. 1996). Two transcripts of 2.7 kb and 7.5 kb are obtained, which are possibly the result of alternative use of distinct polyadenylation sites (Puente et al. 1996). MMP-17 has a furin processing recognition motif, RXR/KR, within its structure (Puente et al. 1996). Further characterization has revealed that functional MMP-17 is translated into proteins of 67 and 71 kDa sizes (Kajita et al. 1999). Human MMP-17 is able to cleave gelatin, but not other typical ECM components (Kolkenbrock et al. 1999, Wang et al. 1999), whereas mouse MMP-17 exhibits the capability of hydrolysing pro-tumour necrosis factor α (TNF-α), fibrinogen and fibrin (English et al. 2000), suggesting differences in functions between species.
Human MMP-25 (MT6-MMP, leukolysin) is a 562 amino acid polypeptide expressed by leukocytes and neutrophils, and in lung, spleen and brain tumours (Velasco et al. 2000, Kang et al. 2001). PMA or IL-8 induces release of MMP-25 form neutrophils as a soluble enzyme with molecular size of 56 kDa and minor forms ranging from 38 to 45 kDa (Kang et al. 2001). MMP-25 degrades type IV collagen, gelatin, fibronectin and fibrin (English et al. 2001).
2.8.8. Type II transmembrane MMPs
MMP-21 and MMP-22 genes were found to be identical to each other. These genes are located on chromosome 1p36.3, which was duplicated during evolution yielding two identical enzymes (Gururajan et al. 1998a, 1998b). Currently, NCBI has named MMP-21 as MMP-23A and MMP-22 as MMP-23B. MMP-23, cloned by Velasco and colleagues (1999), is identical with MMP-23B. A characteristic feature of these genes is that instead of a consensus cysteine-switch motif, they have a sequence ALCLLPA containing a single conserved cysteine for maintaining the enzyme latency. The open reading frame of MMP-23B codes for a protein of 390 amino acids with a molecular mass of 43.9 kDa (Velasco et al. 1999). The MMP-23B protein contains four potential N-glycosylation sites. Structural analysis of the mouse counterpart of MMP-23 has revealed that it has an N-terminal signal anchor targeting it to the cell membrane (Pei et al. 2000). MMP-23 also has cysteine array and immunoglobulin (Ig)-like domains in its C-terminal end (Pei et al. 2000). Several transcripts are expressed due to alternative splicing of MMP-23A and MMP-23B, and these are expressed in heart, placenta, ovary, testis and prostate (Gururajan et al. 1998a, Velasco et al. 1999).