Development of laminin receptor agonists: identification of important functional residues by alanine scanning

An antagonist of cellular adhesion and motility, acetyl-C-S-Acm]-VIGYSGDRC-S-Acm]-NH(2) (mEGF(33-42)), shares homology with the agonist sequence CDPGYIGSR-NH(2). It has been proposed that the latter peptide binds to the high affinity 67 kDa laminin receptor. Both peptides have equal affinities for the receptor and similar conformations have been derived for both. We have examined the importance of individual non-homologous residues with respect to receptor binding and antagonistic properties of mEGF(33-42). Alanine scanning of non-conserved residues in the N-terminal half of mEGF(33-42) caused loss of biological activity with respect to cell attachment, receptor binding and migratory response. Substitution of alanine for serine (position 6) caused loss of laminin-specific cell attachment and receptor binding activities. However, the peptide did stimulate migration suggesting that this peptide may be a non-specific stimulator of migration. In contrast, alanine substitution for the C-terminal Cys-S-Acm had no apparent effect on the attachment or receptor binding activities of the peptide but generated an agonist from the antagonist parent. Comparison of the modelled folds of the alanine containing peptides revealed the presence of significant helical content in those peptides capable of stimulating migration and suggests that a reduction in bulk in the N-terminal residues is not conducive to adopting a productive binding conformation.