Structural and functional analysis of the globular domain IVa of the laminin alpha 1 chain and its impact on an adjacent RGD site.
The globular domain IVa (about 250 residues) of the laminin alpha1 chain was obtained in recombinant form from mammalian cell clones. It was prepared either with (alpha1IVa-R) or without (alpha1IVa) an adjacent cell-adhesive RGD site which seems to be masked in laminin-1. The recombinant products could be visualized as globular structures by rotary shadowing, were resistant to trypsin and shared immunological epitopes with laminin-1, indicating folding into a native structure. Sequence analysis of pepsin fragments demonstrated the insertion of the globular domain into an epidermal growth factor-like scaffold which is characteristic of the extracellular laminin domain IV (L4) module. Only little immunological cross-reaction was found, however, with other L4 modules from perlecan and different laminin isoforms. Fragment alpha1IVa-R, but not fragment alpha1IVa, bound to alphaVbeta3 integrin, although to a distinctly lower level than a laminin fragment where the RGD site is fully exposed. The fragments also had no or only little cell attachment activity. This confirmed previous predictions that the globular domain alpha 1IVa masks the RDG site in laminin-1. Domain alpha 1IVa showed, in addition, a weak binding activity for the basement-membrane protein fibulin-1.
Schulze
B
B
Max-Planck-Institut für Biochemie, Martinsried, Germany.
Mann
K
K
Poschl
E
E
Yamada
Y
Y
Timpl
R
R
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002448:Cell Adhesion; D002999:Clone Cells; D003001:Cloning, Molecular; D017931:DNA Primers; D005260:Female; D006801:Humans; D016023:Integrins / Q000737:chemistry / Q000302:isolation & purification; D007797:Laminin / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D008024:Ligands; D046911:Macromolecular Substances; D008322:Mammals; D008545:Melanoma; D008969:Molecular Sequence Data; D009842:Oligopeptides; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010920:Placenta; D011247:Pregnancy; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D016023:Integrins; D007797:Laminin; D008024:Ligands; D046911:Macromolecular Substances; D009842:Oligopeptides; D010446:Peptide Fragments; D011994:Recombinant Proteins; C047981:arginyl-glycyl-aspartic acid