Mapping the ligand-binding pocket of integrin α5β1 using a gain-of-function approach
- 11 November 2009
- journal article
- research article
- Published by Portland Press Ltd. in Biochemical Journal
- Vol. 424 (2), 179-189
- https://doi.org/10.1042/bj20090992
Abstract
Integrin α5β1 is a key receptor for the extracellular matrix protein fibronectin. Antagonists of human integrin α5β1 have therapeutic potential as anti-angiogenic agents in cancer and diseases of the eye. However, the structure of the integrin is unsolved and the atomic basis of fibronectin and antagonist binding by integrin α5β1 is poorly understood. In the present study, we demonstrate that zebrafish α5β1 integrins do not interact with human fibronectin or the human α5β1 antagonists JSM6427 and cyclic peptide CRRETAWAC. Zebrafish α5β1 integrins do bind zebrafish fibronectin-1, and mutagenesis of residues on the upper surface and side of the zebrafish α5 subunit β-propeller domain shows that these residues are important for the recognition of the Arg-Gly-Asp (RGD) motif and the synergy sequence [Pro-His-Ser-Arg-Asn (PHSRN)] in fibronectin. Using a gain-of-function analysis involving swapping regions of the zebrafish integrin α5 subunit with the corresponding regions of human α5 we show that blades 1–4 of the β-propeller are required for human fibronectin recognition, suggesting that fibronectin binding involves a broad interface on the side and upper face of the β-propeller domain. We find that the loop connecting blades 2 and 3 of the β-propeller, the D3–A3 loop, contains residues critical for antagonist recognition, with a minor role played by residues in neighbouring loops. A new homology model of human integrin α5β1 supports an important function for D3–A3 loop residues Trp157 and Ala158 in the binding of antagonists. These results will aid the development of reagents that block integrin α5β1 functions in vivo.Keywords
This publication has 53 references indexed in Scilit:
- Structural and therapeutic insights from the species specificity and in vivo antithrombotic activity of a novel αIIb-specific αIIbβ3 antagonistBlood, 2009
- Automated image‐based phenotypic analysis in zebrafish embryosDevelopmental Dynamics, 2009
- Species differences in small molecule binding to αIIbβ3 are the result of sequence differences in 2 loops of the αIIb β propellerBlood, 2009
- Linking integrin conformation to functionJournal of Cell Science, 2009
- Volociximab, a Chimeric Monoclonal Antibody that Specifically Binds α5β1 Integrin: A Phase I, Pharmacokinetic, and Biological Correlative StudyClinical Cancer Research, 2008
- Structural basis for distinctive recognition of fibrinogen γC peptide by the platelet integrin αIIbβ3The Journal of cell biology, 2008
- Loss of E-Cadherin Promotes Ovarian Cancer Metastasis via α5-Integrin, which Is a Therapeutic TargetCancer Research, 2008
- In vivo drug discovery in the zebrafishNature Reviews Drug Discovery, 2005
- An Integrin-Dependent Role of Pouch Endoderm in Hyoid Cartilage DevelopmentPLoS Biology, 2004
- IntegrinsCell, 2002