Mapping of potent and specific binding motifs, GLOGEN and GVOGEA, for integrin ?1?1 using collagen toolkits II and III.

ABSTRACT: Integrins are well characterized cell surface receptors for extracellular matrix proteins. Mapping integrin-binding sites within the fibrillar collagens identified GFOGER as a high affinity site recognized by ?2?1, but with lower affinity for ?1?1. Here, to identify specific ligands for ?1?1, we examined binding of the recombinant human ?1 I domain, the rat pheochromocytoma cell line (PC12), and the rat glioma Rugli cell line to our collagen Toolkit II and III peptides using solid-phase and real-time label-free adhesion assays. We observed Mg(2+)-dependent binding of the ?1 I domain to the peptides in the following rank order: III-7 (GLOGEN), II-28 (GFOGER), II-7 and II-8 (GLOGER), II-18 (GAOGER), III-4 (GROGER). PC12 cells showed a similar profile. Using antibody blockade, we confirmed that binding of PC12 cells to peptide III-7 was mediated by integrin ?1?1. We also identified a new ?1?1-binding activity within peptide II-27. The sequence GVOGEA bound weakly to PC12 cells and strongly to activated Rugli cells or to an activated ?1 I domain, but not to the ?2 I domain or to C2C12 cells expressing ?2?1 or ?11?1. Thus, GVOGEA is specific for ?1?1. Although recognized by both ?2?1 and ?11?1, GLOGEN is a better ligand for ?1?1 compared with GFOGER. Finally, using biosensor assays, we show that although GLOGEN is able to compete for the ?1 I domain from collagen IV (IC(50) ?3 ?m), GFOGER is much less potent (IC(50) ?90 ?m), as shown previously. These data confirm the selectivity of GFOGER for ?2?1 and establish GLOGEN as a high affinity site for ?1?1.

SUBMITTER: Hamaia SW 

PROVIDER: S-EPMC3406685 | biostudies-literature | 2012 Jul

REPOSITORIES: biostudies-literature