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

M. Kreiner, Z. Li, J. Beattie, S.M. Kelly, H.J. Mardon, C.F. van der Walle

Research output: Contribution to journalArticle

18 Citations (Scopus)

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.
LanguageEnglish
Pages553-560
Number of pages7
JournalProtein Engineering Design and Selection
Volume21
Issue number9
DOIs
Publication statusPublished - 2008

Fingerprint

Cell adhesion
Integrins
Ligands
Fibronectins
Cell Adhesion
Display devices
Arginine
Size exclusion chromatography
Fasteners
Dichroism
Surface plasmon resonance
Substrates
Hinges
Fibroblasts
Leucine Zippers
Dimers
Streptavidin
Amino acids
Biotinylation
Sulfides

Keywords

  • cell adhesion
  • coiled coil
  • fibronectin
  • integrin
  • polyvalency
  • pharmacology

Cite this

Kreiner, M. ; Li, Z. ; Beattie, J. ; Kelly, S.M. ; Mardon, H.J. ; van der Walle, C.F. / Self-assembling multimeric integrin 5β1 ligands for cell attachment and spreading. In: Protein Engineering Design and Selection. 2008 ; Vol. 21, No. 9. pp. 553-560.
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Self-assembling multimeric integrin 5β1 ligands for cell attachment and spreading. / Kreiner, M.; Li, Z.; Beattie, J.; Kelly, S.M.; Mardon, H.J.; van der Walle, C.F.

In: Protein Engineering Design and Selection, Vol. 21, No. 9, 2008, p. 553-560.

Research output: Contribution to journalArticle

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AU - Li, Z.

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AU - Kelly, S.M.

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AU - van der Walle, C.F.

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