Laminin gamma3 chain binds to nidogen and is located in murine basement membranes.

Recently a novel laminin gamma3 chain was identified in mouse and human and shown to have the same modular structure as the laminin gamma1 chain. We expressed two fragments of the gamma3 chain in mammalian cells recombinantly. The first, domain VI/V, consisting of laminin N-terminal (domain VI) and four laminin-type epidermal growth factor-like (domain V) and laminin N-terminal modules, was shown to be essential for self-assembly of laminins.

Ultrastructural colocalization of nidogen-1 and nidogen-2 with laminin-1 in murine kidney basement membranes.

Nidogen-1, a key component of basement membranes, is considered to function as a link between laminin and collagen type IV networks. Recently a new member of the nidogen family, nidogen-2, has been characterized. Preliminary immunohistochemical data indicated that nidogen-1 and nidogen-2 show a similar tissue distribution at the light microscopic level.

Recombinant domain IV of perlecan binds to nidogens, laminin-nidogen complex, fibronectin, fibulin-2 and heparin.

Domain IV of mouse perlecan, which consists of 14 immunoglobulin superfamily (IG) modules, was prepared from recombinant human cell culture medium in the form of two fragments, IV-1 (IG2-9, 100 kDa) and IV-2 (IG10-15, 66 kDa). Both fragments bound to a heparin column, being eluted at ionic strengths either below (IV-2) or above (IV-1) physiological level, and could thus be readily purified. Electron microscopy demonstrated an elongated shape (20-25 nm), and folding into a native structure was indicated by immunological assay and CD spectroscopy.

Structural and genetic analysis of laminin-nidogen interaction.

High-affinity binding of nidogen to laminins involves a single binding site on the laminin gamma 1 chain and is thus a property shared by almost all laminin isoforms. This binding mediates the connection of laminins to the collagen IV network, perlecan and other proteins and is considered to be an essential step in the stabilization of basement membranes. Nidogen binding has been located to a single LE module (gamma 1III4) by recombinant analysis.

Nidogen-2: a new basement membrane protein with diverse binding properties.

Human nidogen-2 was cloned and sequenced (1375 residues) and found to share 46% sequence identity and a similar domain arrangement with the previously characterized basement membrane protein nidogen-1. Recombinant nidogen-2 was purified as a 200 kDa protein from transfected mammalian cell medium, showed a high level of N and O-glycosylation, and could be clearly distinguished from nidogen-1 (150 kDa) by specific antibodies. Electron microscopy demonstrated that the two isoforms have a similar shape, consisting of three globular domains connected by two threads, but differ somewhat in length.