Amino acid changes in Drosophila αPS2βPS integrins that affect ligand affinity

Thomas A. Bunch, Teresa L. Helsten, Timmy L. Kendall, Nikhil Shirahatti, Daruka Mahadevan, Sanford J. Shattil, Danny L. Brower

Research output: Contribution to journalArticlepeer-review

2 Scopus citations


We developed a ligand-mimetic antibody Fab fragment specific for Drosophila αPS2βPS integrins to probe the ligand binding affinities of these invertebrate receptors. TWOW-1 was constructed by inserting a fragment of the extracellular matrix protein Tiggrin into the H-CDR3 of the αvβ3 ligand-mimetic antibody WOW-1. The specificity of αPS2βPS binding to TWOW-1 was demonstrated by numerous tests used for other integrin-ligand interactions. Binding was decreased in the presence of EDTA or RGD peptides and by mutation of the TWOW-1 RGD sequence or the βPS metal ion-dependent adhesion site (MIDAS) motif. TWOW-1 binding was increased by mutations in the αPS2 membrane-proximal cytoplasmic GFFNR sequence or by exposure to Mn2+. Although Mn2+ is sometimes assumed to promote maximal integrin activity, TWOW-1 binding in Mn2+ could be increased further by the αPS2 GFFNR→GFANA mutation. A mutation in the βPS I domain (βPS-b58; V409D) greatly increased ligand binding affinity, explaining the increased cell spreading mediated by αPS2βPS-b58. Further mutagenesis of this residue suggested that Val-409 normally stabilizes the closed head conformation. Mutations that potentially reduce interaction of the integrin β subunit plexin-semaphorin-integrin (PSI) and stalk domains have been shown to have activating properties. We found that complete deletion of the βPS PSI domain enhanced TWOW-1 binding. Moreover the PSI domain is dispensable for at least some other integrin functions because βPS-ΔPSI displayed an enhanced ability to mediate cell spreading. These studies establish a means to evaluate mechanisms and consequences of integrin affinity modulation in a tractable model genetic system.

Original languageEnglish (US)
Pages (from-to)5050-5057
Number of pages8
JournalJournal of Biological Chemistry
Issue number8
StatePublished - Feb 24 2006
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


Dive into the research topics of 'Amino acid changes in Drosophila αPS2βPS integrins that affect ligand affinity'. Together they form a unique fingerprint.

Cite this