Unraveling the amino acid sequence crucial for heparin binding to collagen V.
We have previously shown that a recombinant 12-kDa fragment of the collagen alpha1(V) chain (Ile(824)-Pro(950)), referred to as HepV, binds to heparin and heparan sulfate (Delacoux, F., Fichard, A., Geourjon, C., Garrone, R., and Ruggiero, F. (1998) J. Biol. Chem. 273, 15069-15076). No consensus sequence was found in the alpha1(V) primary sequence, but a cluster of 7 basic amino acids (in the Arg(900)-Arg(924) region) was postulated to contain the heparin-binding site. The contribution of individual basic amino acids within this sequence was examined by site-directed mutagenesis. Further evidence for the precise localization of the heparin-binding site was provided by experiments based on the fact that heparin can protect the alpha1(V) chain heparin-binding site from trypsin digestion. The results parallel the alanine scanning mutagenesis data, i.e. heparin binding to the alpha1(V) chain involved Arg(912), Arg(918), and Arg(921) and two additional neighboring basic residues, Lys(905) and Arg(909). Our data suggest that this extended sequence functions as a heparin-binding site in both collagens V and XI, indicating that these collagens use a novel sequence motif to interact with heparin.
Delacoux
F
F
Institut de Biologie et Chimie des Protéines, CNRS UPR 412, Université Claude Bernard, 7, Passage du Vercors, 69367 Lyon Cedex 07, France.
Fichard
A
A
Cogne
S
S
Garrone
R
R
Ruggiero
F
F
D000595:Amino Acid Sequence; D000596:Amino Acids; D001483:Base Sequence; D001665:Binding Sites; D003094:Collagen / Q000737:chemistry* / Q000378:metabolism; D006493:Heparin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding
D000596:Amino Acids; D006493:Heparin; D003094:Collagen