Matrix Metalloproteinases, Vascular Remodeling, and Vascular Disease

Abstract

Matrix metalloproteinases (MMPs) are a family of zinc-dependent endopeptidases that degrade various proteins in the extracellular matrix (ECM). Typically, MMPs have a propeptide sequence, a catalytic metalloproteinase domain with catalytic zinc, a hinge region or linker peptide, and a hemopexin domain. MMPs are commonly classified on the basis of their substrates and the organization of their structural domains into collagenases, gelatinases, stromelysins, matrilysins, membrane-type (MT)-MMPs, and other MMPs. MMPs are secreted by many cells including fibroblasts, vascular smooth muscle (VSM), and leukocytes. MMPs are regulated at the level of mRNA expression and by activation through removal of the propeptide domain from their latent zymogen form. MMPs are often secreted in an inactive proMMP form, which is cleaved to the active form by various proteinases including other MMPs. MMPs degrade various protein substrates in ECM including collagen and elastin. MMPs could also influence endothelial cell function as well as VSM cell migration, proliferation, Ca²⁺ signaling, and contraction. MMPs play a role in vascular tissue remodeling during various biological processes such as angiogenesis, embryogenesis, morphogenesis, and wound repair. Alterations in specific MMPs could influence arterial remodeling and lead to various pathological disorders such as hypertension, preeclampsia, atherosclerosis, aneurysm formation, as well as excessive venous dilation and lower extremity venous disease. MMPs are often regulated by endogenous tissue inhibitors of metalloproteinases (TIMPs), and the MMP/TIMP ratio often determines the extent of ECM protein degradation and tissue remodeling. MMPs may serve as biomarkers and potential therapeutic targets for certain vascular disorders.

Keywords: Aneurysm; Angiogenesis; Atherosclerosis; Cell signaling; Extracellular matrix; Hypertension; Smooth muscle; TIMPs.

Fig. 1

MMP subtypes and their structure. A typical MMP consists of a propeptide, a catalytic metalloproteinase domain, a linker peptide (hinge region), and a hemopexin domain. The propeptide has a cysteine switch PRCGXPD whose cysteine sulfhydryl (–SH) group chelates the active site Zn²⁺, and keeps the MMP in its latent proMMP zymogen form. The catalytic domain contains the Zn²⁺ binding motif HEXXHXXGXXH, two Zn²⁺ ions (one catalytic and one structural), specific S1, S2,…Sn and S1’, S2’,…Sn’ pockets, which confer specificity, and two or three Ca²⁺ ions for stabilization. Some MMPs show exceptions in their structures. Gelatinases have 3 type-II fibronectin repeats in the catalytic domain. Matrilysins have neither a hinge region nor a hemopexin domain. Furin-containing MMPs such as MMP-11, 21 and 28 have a furin-like pro-protein convertase recognition sequence in the propeptide C-terminus. MMP-28 has a slightly different cysteine switch motif PRCGVTD. Membrane-type MMPs (MT-MMPs) typically have a transmembrane domain and a cytosolic domain. MMP-17 and -25 have a glycosylphosphatidylinositol (GPI) anchor. MMP-23 lacks the consensus PRCGXPD motif, has a cysteine residue located in a different sequence ALCLLPA, may remain in the latent inactive proform through its type-II signal anchor, and has a cysteine-rich region and an immunoglobulin-like proline-rich region.

Fig. 2

MMP-substrate interaction. MMP-3 is used as an example, and the MMP-substrate-interaction and the positions of the conserved His and Glu may vary in other MMPs. Only the MMP catalytic domain is illustrated, and the remaining part of the MMP molecule is truncated by squiggles. A) In the quiescent MMP molecule, the catalytic Zn²⁺ is supported in the HEXXHXXGXXH-motif by binding to the imidazole rings of the 3 histidines His201, 205, 211. Additionally, the methionine-219 (Met219) in the conserved XBMX Met-turn acts as a hydrophobic base to further support the structure surrounding the catalytic Zn²⁺. In preparation for substrate binding, an incoming H₂O molecule is polarized between the MMP acidic Zn²⁺ and basic glutamate-202 (Glu202). B) Using H⁺ from free H₂O, the substrate carbonyl group binds to Zn²⁺, forming a Michaelis complex. This allows the MMP S1, S2, S3, …Sn pockets on the right side of Zn²⁺ and the primed S1’, S2’, S3’, …Sn’ pockets on the left side of Zn²⁺ to confer specific binding to the substrate P1, P2, P3, … Pn and the primed P1’, P2’, P3’, … Pn’ substituents, respectively. The MMP pockets are organized such that the S1 and S3 pockets are located away from the catalytic Zn²⁺, while the S2 pocket is closer to Zn²⁺. C) The substrate-bound H₂O is freed, the Zn²⁺-bound oxygen from the Glu-bound H₂O executes a nucleophilic attack on the substrate carbon, and the Glu202 extracts a proton from the Glu-bound H₂O to form an N-H bond with the substrate N, resulting in a tetrahedral intermediate. D) Freed H₂O is taken up again, and the second proton from Glu-bound H₂O is transferred to the substrate, forming an additional N-H bond. As a result, the substrate scissile C-N bond breaks, thus releasing the N portion of the substrate while the carboxylate portion of the substrate remains in an MMP-carboxylate complex. Another H₂O is taken up, thus releasing the remaining carboxylate portion of the substrate, and the MMP is prepared to attack another substrate (A).

Fig. 3

TIMP-MMP Interaction. TIMP-1 and MMP-3 are used as prototypes. The amino acids involved in Zn²⁺- and pocket-binding may vary with different MMPs and TIMPs. (A) TIMP is a ~190 aa protein, with an N-terminal domain (loops L1, L2, and L3) and C-terminal domain (loops L4, L5 and L6), which fold independently as a result of 6 disulfide bonds between 12 specific Cys residues. The N-terminal Cys1-Thr-Cys-Val4 and Glu67-Ser-Val-Cys70 are connected via a disulfide bond between Cys1 and Cys70 and are essential for MMP inhibition, as they enter the MMP active site and bidentately chelate the MMP Zn²⁺. The carbonyl oxygen and α-amino nitrogen in the TIMP Cys1 coordinate with the MMP Zn²⁺, which is localized in the MMP molecule via the 3 histidines in the HEXXHXXGXXH motif. The TIMP α-amino group then expels Zn²⁺-bound H₂O by binding the MMP H₂O binding site and forming a hydrogen bond with carboxylate oxygen from conserved MMP Glu202 (E in the HEXXHXXGXXH sequence). (B) TIMP Thr2 side chains enter the MMP S1’ pocket in a manner similar to that of a substrate P1’ substituent. Thr2 –OH group could also interact with Glu202, further contributing to expelling Zn²⁺-bound H₂O and preventing substrate degradation. The TIMP Cys3, Val4 and Pro5 also interact with MMP S2′, S3’, and S4′ pockets in a P2′, P3′, and P4’-like manner, further preventing substrate binding.

Fig. 4

Representative roles of MMPs in vascular biology.

Fig. 5

Representative roles of MMPs in vascular pathology.