A single EGF-like motif of laminin is responsible for high affinity nidogen binding

Mayer, U ORCID: https://orcid.org/0000-0003-2328-0052, Nischt, R, Pöschl, E, Mann, K, Fukuda, K, Gerl, M, Yamada, Y and Timpl, R (1993) A single EGF-like motif of laminin is responsible for high affinity nidogen binding. The EMBO Journal, 12 (5). pp. 1879-1885. ISSN 0261-4189

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A major nidogen binding site of mouse laminin was previously localized to about three EGF-like repeats (Nos 3-5) of its B2 chain domain III [M. Gerl et al. (1991) Eur. J. Biochem., 202, 167]. The corresponding cDNA was amplified by polymerase chain reaction and inserted into a eukaryotic expression vector tagged with a signal peptide. Stably transfected human kidney cell clones were shown to process and secrete the resulting fragment B2III3-5 in substantial quantities. It possessed high binding activity for recombinant nidogen in ligand assays, with an affinity comparable with that of authentic laminin fragments. In addition, complexes of B2III3-5 and nidogen could be efficiently converted into a covalent complex by cross-linking reagents. Proteolytic degradation of the covalent complex demonstrated the association of B2III3-5 with a approximately 80 residue segment of nidogen domain G3 to which laminin binding has previously been attributed. The correct formation of most of the 12 disulfide bridges in B2III3-5 was indicated from its protease resistance and the complete loss of cross-reacting epitopes as well as of nidogen-binding activity after reduction and alkylation. Smaller fragments were prepared by the same recombinant procedure and showed that combinations of EGF-like repeats 3-4 and 4-5 and the single repeat 4 but not repeats 3 or 5 possess full nidogen-binding activity. This identifies repeat 4 as the only binding structure. The sequence of repeat 4 is well conserved in the human and in part in the Drosophila laminin B2 chain.(ABSTRACT TRUNCATED AT 250 WORDS)

Item Type: Article
Uncontrolled Keywords: amino acid sequence,animals,antibodies,base sequence,binding sites,cross-linking reagents,epidermal growth factor,genetic vectors,laminin,membrane glycoproteins,mice,molecular sequence data,oligodeoxyribonucleotides,peptide fragments,protein binding,protein conformation,recombinant proteins,transfection
Faculty \ School: Faculty of Science > School of Biological Sciences
UEA Research Groups: Faculty of Science > Research Groups > Cells and Tissues
Faculty of Medicine and Health Sciences > Research Groups > Gastroenterology and Gut Biology
Depositing User: Pure Connector
Date Deposited: 12 Jan 2016 17:00
Last Modified: 19 Apr 2023 23:45
URI: https://ueaeprints.uea.ac.uk/id/eprint/56270
DOI: 10.1002/j.1460-2075.1993.tb05836.x

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