Two non-contiguous regions contribute to nidogen binding to a single EGF-like motif of the laminin .gamma.1 chain

US5493008A · kind A · utility

Assignee

• University of Virginia Patent Foundation · US

Inventors

• Jay W. Fox · Charlottesville, US
• Rupert Timpl · Gauting, DE

Key dates

• Filing date: Aug 15, 1994
• Grant date: Feb 20, 1996
• Expiry date: Aug 15, 2014

Classification

• Technology area (CPC A): Human Necessities
• CPC primary: A61K38/00
• WIPO field: Pharmaceuticals
• WIPO sector: Chemistry

Abstract

High affinity binding of nidogen to laminin is mediated by an EGF-like repeat .gamma.1III4 of the mouse laminin .gamma.1 chain and has now been restricted to two short non-contiguous regions of its 56 residue sequence by use of synthetic peptides and recombinant mutants. Disulfide loop a,b of the repeat and a modified loop a,c could completely inhibit binding, with a 5,000-fold or 300-fold reduced affinity, respectively. Synthetic loops c and d lacked inhibitory activity. Some binding contribution of Try819 in loop c was, however, shown by mutation and side chain modification. Together with studies of loop chimeras, this indicated a distinct cooperativity between the two binding sites. The major binding site of loop a was localized to the heptapeptide NIDPNAV (position 798-804). A change of Asp800 to Asn or Ala803 to Val caused a strong reduction in binding activity, while only small effects were observed for the changes Pro801 to Gln and Ile799 to Val. The latter replacement corresponds to the single substitution found in the same region of the Drosophila laminin .gamma.1 chain. However, the changes Asn802 to Ser or Val804 to Ser, both known to exist in the laminin .gamma.2 chain,…