Laminins, heterotrimeric glycoproteins in the basement membrane, are involved in diversebiological activities. So far, five
, three
, and three
chains have been identified, and at least 15laminin isoforms exist composed of various combinations of the different three chains. The major cell-surface receptors for laminins are integrins and proteoglycans, such as dystroglycans and syndecans.Previously, we reported that synthetic peptide A4G82 (TLFLAHGRLVFM, mouse laminin
4 chainresidues 1514-1525) showed strong cell attachment and syndecan binding activities. On the basis of thecrystal structure of the LG module and sequence alignment, A4G82 is located in the connecting loopregion between
-strands E and F in the laminin
4 chain LG4 module. Here, we have focused on thestructural importance of this E-F loop region for the biological activity of the
4 chain LG4 module. Todetermine the importance of the loop structure, we synthesized peptide A4G82X (
cyclo-A4G82X, Cys-TLFLAHGRLVFX-Cys, X= norleucine), which was cyclized via disulfide bridges at both the N- andC-termini. The cyclic peptides derived from A4G82X inhibited the heparin binding activity of the
4chain G domain and promoted HT-1080 cell attachment better than the corresponding linear peptides.We determined FLAHGRLVFX as a minimal sequence of
cyclo-A4G82X important for cell adhesionand heparin binding using a series of truncated peptides. Moreover, HT-1080 cell attachment to the cyclicpeptides was more efficiently blocked by heparin than cell attachment to the linear peptides. Furthermore,the cyclic peptides showed significantly enhanced syndecan-2-mediated cell attachment activity. Theseresults indicate that the activity of A4G82 is highly conformation-dependent, suggesting that the E-Floop structure is crucial for its biological activity.
