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The Strathprints institutional repository is a digital archive of University of Strathclyde research outputs.

Strathprints serves world leading Open Access research by the University of Strathclyde, including research by the Strathclyde Institute of Pharmacy and Biomedical Sciences (SIPBS), where research centres such as the Industrial Biotechnology Innovation Centre (IBioIC), the Cancer Research UK Formulation Unit, SeaBioTech and the Centre for Biophotonics are based.

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Self-assembling multimeric integrin 5β1 ligands for cell attachment and spreading

Kreiner, M. and Li, Z. and Beattie, J. and Kelly, S.M. and Mardon, H.J. and van der Walle, C.F. (2008) Self-assembling multimeric integrin 5β1 ligands for cell attachment and spreading. Protein Engineering Design and Selection, 21 (9). pp. 553-560. ISSN 1741-0126

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Abstract

Substrates utilising clustered arginine-glycine-aspartic acid (RGD) ligand displays support greater cell adhesion over random displays. However, cell adhesion to integrin 5β1 requires the synergy site on the 9th type III fibronectin domain (FIII) in addition to RGD on the 10th FIII domain. Here, we have designed and expressed soluble protein chimeras consisting of an N-terminal 9th-10th FIII domain pair, IgG-derived hinge and leucine zipper-derived helix; the latter mutated to yield di-, tri- and tetrameric coiled coils and thus self-assembling, multimeric integrin 5β1 ligands. A unique C-terminal cysteine was appended to the helix to facilitate 'anchoring' of the chimeras with a defined orientation on a surface. Size-exclusion chromatography and circular dichroism demonstrated that the chimeras self-assembled as multimers in solution with defined secondary structures predicted from theoretical calculations. Biotinylation via a thioether bond was used to selectively bind the chimeras to streptavidin-coated surfaces, each of which was then shown to bind integrin 5β1 by surface plasmon resonance. Spreading of fibroblasts to surfaces derivatised with the chimeras was found to proceed in the order: tetramer > trimer > dimer > monomer. Thus, we describe novel polyvalent integrin 5β1 ligands for facile derivatisation of substrates to improve cell adhesion in vitro.