The Type XIII Collagen Ectodomain Is a 150-nm Rod and Capable of Binding to Fibronectin, Nidogen-2, Perlecan, and Heparin

Abstract

Type XIII collagen consists of a short N-terminal intracellular domain, a transmembrane domain, and a collagenous ectodomain, and it is found at many sites of cell adhesion. We report on the characterization of recombinant type XIII collagen. The shed ectodomain was purified from insect cell culture medium and shown to form 240-kDa trimers with a T m of 42°C. Correct chain association into a triple-helical conformation was confirmed by limited pepsin digestion and CD spectroscopy. Rotary shadowing electron microscopy of the ectodomain revealed it to be a 150-nm rod with two flexible hinges separating 31-, 52-, and 68-nm portions. The rods represent the collagenous domains 1-3, and the hinges coincide with the non-collagenous domains 2 and 3. By using surface plasmon resonance analysis, the ectodomain showed interaction with immobilized fibronectin, nidogen-2, and perlecan with K D values in the nanomolar range. The binding sites of type XIII collagen for fibronectin were localized to the collagenous domains, whereas the binding activities for nidogen-2 and perlecan resided in the pepsin-sensitive portions of the ectodomain. Furthermore, the ectodomain bound significantly to heparin, which also inhibited shedding of the ectodomain in insect cell cultures. The results reveal that type XIII collagen is notably distinct in its structure compared with other cell-surface proteins, and the in vitro binding with fibronectin, heparin, and two basement membrane components is indicative of multiple cell-matrix interactions in which this ubiquitously expressed protein participates.Two of the 19 collagens described in vertebrates belong to the subgroup of transmembrane collagens, namely type XIII and the hemidesmosomal component, type XVII collagen (1-3). These two nonfibrillar collagens are not structurally homologous except that both have a transmembrane domain. The collagen superfamily also includes other transmembrane proteins that have short collagenous domains, namely the types I and II macrophage scavenger receptors; C1q, the first component of complement C1; a macrophage receptor with a collagenous structure; and the ectodysplasin-A family of proteins (1, 3, 4).Type XIII collagen molecules reside in the plasma membrane of cells in a type II orientation with a short N-terminal cytosolic domain, a single transmembrane domain, and a large, mainly collagenous ectodomain (5, 6). The N-terminal non-collagenous domain, NC1, 1 of the human protein encompasses a 38-residue cytosolic domain, a 23-residue transmembrane domain, and the first 60 residues of the non-collagenous extracellular sequences adjacent to the plasma membrane (5). The rest of the ectodomain contains three collagenous sequences, COL1-3, with sizes of 104, 172, and 235 residues, respectively, and non-collagenous domains, NC2-4, with sizes of 34, 22, and 18 residues (5). The precursor RNAs encoding human and mouse type XIII collagen are known to be subject to complex alternative splicing, and hence the sizes of the COL1, NC2, COL3, and NC4 domains in humans ca...

WARP Is a Novel Multimeric Component of the Chondrocyte Pericellular Matrix That Interacts with Perlecan

“…The perlecan core protein has been shown to associate with a range of basement membrane macromolecules including collagens (IV and XIII), laminin-1, nidogens 1 and 2, ␤1-integrin, ␣-dystroglycan, as well as fibulin-2, fibrillin-1, ECM-1, PRELP (proline-arginine-rich end leucine-rich repeat protein), fibronectin, heparin, and several growth factors (platelet-derived growth factor, FGF-2, FGF-7) and FGF-binding protein (19,(32)(33)(34)(35)(36)(37).…”

Type XIX Collagen Purified from Human Umbilical Cord Is Characterized by Multiple Sharp Kinks Delineating Collagenous Subdomains and by Intermolecular Aggregates via Globular, Disulfide-linked, and Heparin-binding Amino Termini

“…10 and references therein and Refs. [45][46][47]. The kinks/bends that have been observed are often difficult to map to specific interruptions when the collagenous subdomains are not as defined as they are in type XIX and related family members (see Introduction).…”

Recent advances in annular pathobiology provide insights into rim-lesion mediated intervertebral disc degeneration and potential new approaches to annular repair strategies

“…The low density lipoprotein (LDL) repeats in perlecan domain II bind connective tissue growth factor (CTGF) [2,101], CTGF modulates bone morphogenetic protein (BMP) and transforming growth factor (TGF)-b signalling to co-ordinate morphogenesis, and chondrogenic and angiogenic events during skeletal development [2,101]. Perlecan also interacts with a number of matrix stabilising molecules including fibrillin-1, nidogen-1 and 2, fibulin-2, fibronectin, proline/argininerich and leucine-rich repeat protein/prolargin (PRELP), von Willebrand factor protein-A related protein (WARP) and types XIII and XVIII collagen; and cell attachment proteins such as laminin, fibronectin and thrombospondin [6,8,45,54,78,87,141] and thus also has important roles to play in matrix organisation and stabilisation.…”