Interdomain Flexibility in Full-length Matrix Metalloproteinase-1 (MMP-1)

Abstract

The presence of extensive reciprocal conformational freedom between the catalytic and the hemopexin-like domains of fulllength matrix metalloproteinase-1 (MMP-1) is demonstrated by NMR and small angle x-ray scattering experiments. This finding is discussed in relation to the essentiality of the hemopexin-like domain for the collagenolytic activity of MMP-1. The conformational freedom experienced by the present system, having the shortest linker between the two domains, when compared with similar findings on MMP-12 and MMP-9 having longer and the longest linker within the family, respectively, suggests this type of conformational freedom to be a general property of all MMPs. Matrix metalloproteinases (MMP)2 are extracellular hydrolytic enzymes involved in a variety of processes including connective tissue cleavage and remodeling (1-3). All 23 members of the family are able to cleave simple peptides derived from connective tissue components such as collagen, gelatin, elastin, etc. A subset of MMPs is able to hydrolyze more resistant polymeric substrates, such as cross-linked elastin, and partially degraded collagen forms, such as gelatin and type IV collagens (4). Intact triple helical type I-III collagen is only attacked by collagenases MMP-1, MMP-8, and MMP-13 and by MMP-2 and MMP-14 (5-12). Although the detailed mechanism of cleavage of single chain peptides by MMP has been largely elucidated (13-19), little is known about the process of hydrolysis of triple helical collagen. In fact, triple helical collagen cannot be accommodated in the substrate-binding groove of the catalytic site of MMPs (9).All MMPs (but MMP-7) in their active form are constituted by a catalytic domain (CAT) and a hemopexin-like domain (HPX) (20 -22). The CAT domain contains two zinc ions and one to three calcium ions. One zinc ion is at the catalytic site and is responsible for the activity, whereas the other metal ions have structural roles. The isolated CAT domains retain full catalytic activity toward simple peptides and single chain polymeric substrates such as elastin, whereas hydrolysis of triple helical collagen also requires the presence of the HPX domain (9, 23-25). It has been shown that the isolated CAT domain regains a small fraction of the activity of the full-length (FL) protein when high amounts of either inactivated full-length proteins or isolated HPX domains are added to the assay solution (9). Finally, it has been shown that the presence of the HPX domain alone alters the CD spectrum of triple helical collagen in a way that suggests its partial unwinding (26,27). It is tempting to speculate that full-length collagenases attack collagen by first locally unwinding the triple helical structure with the help of the HPX domain and then cleaving the resulting, exposed, single filaments (9, 28).Until 2007, three-dimensional structures of full-length MMPs had been reported only for collagenase MMP-1 (29 -31) and gelatinase MMP-2 (32). The structures of the two proteins are very similar and show a compact arrangement of the two ...

pH dependence of the enzymatic processing of collagen I by MMP-1 (fibroblast collagenase), MMP-2 (gelatinase A), and MMP-14 ectodomain

“…This different mode of controlling the enzymatic activity on the different components of the collagen I triple helix implies that (1) the a-1 chain discriminates among various MMPs through the binding region and/or the binding mode, keeping unchanged the overall tertiary structure of the chain (and thus its susceptibility to cleavage by various MMPs), and (2) the a-2 chain interacts quite similarly with different MMPs, though undergoing different ligand-linked conformational changes, which bring about a MMP-dependent unwinding of the helical arrangement, exposing to a different extent the chain to the proteolytic cleavage. As previously reported, the reciprocal structural flexibility between the hemopexinlike domain and the catalytic domain seems to be a general property of all MMPs [41]. In line with these findings, different levels of conformational freedom seem to be very important, as indicated by different effects on K M (mostly regulated by the interaction with exosites) and on k cat (mostly regulated by the catalytic domain) of the proteolytic processing of different chains of collagen I.…”

Structural insights into triple-helical collagen cleavage by matrix metalloproteinase 1

“…Recent NMR and mutagenesis studies of the Cat domain of MMP-12 characterized several exosites for elastin (48,49). Similar studies were carried out with the isolated Hpx domain of MMP-1 and a triple-helical collagen peptide (40) and suggested a role for Phe282 (Phe301 in the numbering used in ref. 40) in binding of collagen.…”

“…Similar studies were carried out with the isolated Hpx domain of MMP-1 and a triple-helical collagen peptide (40) and suggested a role for Phe282 (Phe301 in the numbering used in ref. 40) in binding of collagen. However, the exact modes of interaction between the enzyme and the target substrate have not been elucidated in these studies.…”

“…Comparison of the crystal structure of proMMP-1 (29) with the structure of the activated form of MMP-1(E200A) (6,7) suggested that the Cat-Hpx linker region confers interdomain flexibility. Such flexing was further demonstrated for MMP-1 by NMR and small angle X-ray scattering studies (40,41) and seems to be a common property of MMPs (42,43). Our crystal structure of MMP-1 bound to collagen has a more closed conformation than free MMP-1 (SI Appendix, Fig.…”

Interstitial Collagen Catabolism

“…MMP-1 is in equilibrium between open/extended and closed structures ( Fig. 2A) (60). An open form of MMP-1 is favored in solution (see below).…”