title: Direct visualization of protease action on collagen triple helical structure.
abstract: Enzymatic processing of extracellular matrix (ECM) macromolecules by matrix metalloproteases (MMPs) is crucial in mediating physiological and pathological cell processes. However, the molecular mechanisms leading to effective physiological enzyme-ECM interactions remain elusive. Only scant information is available on the mode by which matrix proteases degrade ECM substrates. An example is the enzymatic degradation of triple helical collagen II fragments, generated by the collagenase MMP-8 cleavage, during the course of acute inflammatory conditions by gelatinase B/MMP-9. As is the case for many other matrix proteases, it is not clear how MMP-9 recognizes, binds and digests collagen in this important physiological process. We used single molecule imaging to directly visualize this protease during its interaction with collagen fragments. We show that the initial binding is mediated by the diffusion of the protease along the ordered helix on the collagen (3/4) fragment, with preferential binding of the collagen tail. As the reaction progressed and prior to collagen degradation, gelatin-like morphologies resulting from the denaturation of the triple helical collagen were observed. Remarkably, this activity was independent of enzyme proteolysis and was accompanied by significant conformational changes of the working protease. Here we provide the first direct visualization of highly complex mechanisms of macromolecular interactions governing the enzymatic processing of ECM substrates by physiological protease.
authors.affiliation: Department of Structural Biology, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Rosenblum
authors.forename: Philippe E
authors.initials: PE
authors.lastname: Van den Steen
authors.forename: Sidney R
authors.initials: SR
authors.lastname: Cohen
authors.forename: Arkady
authors.initials: A
authors.lastname: Bitler
authors.forename: David D
authors.initials: DD
authors.lastname: Brand
authors.forename: Ghislain
authors.initials: G
authors.lastname: Opdenakker
authors.forename: Irit
authors.initials: I
authors.lastname: Sagi
mesh_terms: D003094:Collagen / Q000737:chemistry* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005109:Extracellular Matrix / Q000378:metabolism; D020782:Matrix Metalloproteinases / Q000378:metabolism*; D018625:Microscopy, Atomic Force; D011487:Protein Conformation
chemical_list: D003094:Collagen; D020782:Matrix Metalloproteinases