Ambidextrous binding of cell and membrane bilayers by soluble matrix metalloproteinase-12

Abstract

Matrix metalloproteinases (MMPs) regulate tissue remodeling, inflammation, and disease progression. Some soluble MMPs are inexplicably active near cell surfaces. Here, we demonstrate binding of MMP-12 directly to bilayers and cellular membranes using paramagnetic NMR and fluorescence. Opposing sides of the catalytic domain engage spin-labeled membrane mimics. Loops project from the β-sheet interface to contact the phospholipid bilayer with basic and hydrophobic residues. The distal membrane interface comprises loops on the other side of the catalytic cleft. Both interfaces mediate MMP-12 association with vesicles and cell membranes. MMP-12 binds plasma membranes and is internalized to hydrophobic perinuclear features, the nuclear membrane, and inside the nucleus within minutes. While binding of TIMP-2 to MMP-12 hinders membrane interactions beside the active site, TIMP-2-inhibited MMP-12 binds vesicles and cells, suggesting compensatory rotation of its membrane approaches. MMP-12 association with diverse cell membranes may target its activities to modulate innate immune responses and inflammation.

Charge-Triggered Membrane Insertion of Matrix Metalloproteinase-7, Supporter of Innate Immunity and Tumors

“…The corresponding but longer MT-loop of MT1-MMP is required for its localization to integrin-containing cell adhesion complexes in plasma membranes where it is enabled to digest collagen fibrils for tumor cell invasion (Woskowicz et al, 2013). The corresponding II-III loop of MMP-12 is the only sequence element that its membrane interfaces (lacking a Trp)(Koppisetti et al, 2014) share with MMP-7. Might other MMPs employ the II-III loop to bind cell membranes as well?…”

“…Paramagnetic structural NMR methods using micelles containing a spin label (Hilty et al, 2004; Kutateladze et al, 2004) have been enhanced and extended to the bilayer phase of disk-like, bilayered micelles known as bicelles (Koppisetti et al, 2014; Lenoir et al, 2015). This approach found MMP-12 to bind bilayer mimics and cellular membranes on opposite sides of the catalytic domain nearer either the β-sheet or α-helices (Koppisetti et al, 2014). ProMMP-7 is reported herein to bind lipid bilayers and mimics near the edges of its β-sheet, but with rather different angles of approach.…”

Multimodality and Molecular Imaging of Matrix Metalloproteinase Activation in Calcific Aortic Valve Disease

“…Activation of these latent MMPs exposes the catalytic domain targeted by 99m Tc-RP805, the tracer used in our studies. Recent data show that activated MMPs bind to, and can be taken up, by neighboring cells ( 21 ). Several MMPs are expressed in CAVD and may contribute to valvular fibrosis and calcification ( 22-24 ).…”

Path to Collagenolysis

“…This reinforces the hypothesis that this loop may be important in the molecular recognition of fibrillar substrates en route to the active site. The II-III loop is also critically important for the molecular recognition of membrane bilayers (39,40) and a receptor (72). These precedents and the PREs from spin-labeled triple-helical peptides encompassing the II-III loop may be consistent with the possibility of a functional role in initial associations with collagen triple helix.…”

“…These 1 H NMR line broadenings are measured as paramagnetic relaxation enhancements (PREs) most reliably as transverse relaxation rate constants, ⌫ 2 (35). The insights from PREs into complexes of lower affinity (35) have been exploited to characterize the structures of mixtures of specific and nonspecific protein complexes with DNA (33), proteins (34), redox protein partners (32,37,38), and bilayer mimics (39,40). Synthesis of a nitroxide-containing artificial amino acid into key locations within a triple-helical peptide model of collagens I-III enabled structural capture of its transient complex with a domain of MT1-MMP (41).…”