Abstract：
:alpha5beta1 integrin can occupy several distinct conformational states which support different strengths of binding to fibronectin [García, A. J., et al. (1998) J. Biol. Chem. 273, 34710-34715]. Using a model system in which specific activating monoclonal antibodies were used to achieve uniform activated states, the binding of alpha5beta1 to full-length wild-type fibronectin and mutants of fibronectin in the defined RGD and PHSRN synergy sites was analyzed using a novel method that measures the strength of the coupling between integrin and its ligand. Neither TS2/16- nor AG89-activated alpha5beta1 showed significant mechanical coupling to RGD-deleted fibronectin. However, peptide competition assays demonstrated a 6-fold difference in the binding affinities of these two states for RGD. The mutant synergy site reduced the AG89 (low)-activated state to background levels, but the TS2/16-activated state still retained approximately 30% of the wild-type activity. Thus, these two active binding states of alpha5beta1 interact differently with both the RGD and synergy domains. The failure of the AG89-activated state to show mechanical coupling to either the RGD or synergy domain mutants was unexpected and implies that the RGD domain itself does not contribute significant mechanical strength to the alpha5beta1-fibronectin interaction. The lack of RGD alone to support alpha5beta1 coupling was further confirmed using a synthetic polymer presenting multiple copies of the RGD loop. These results suggest a model in which the RGD domain serves to activate and align the alpha5beta1-fibronectin interface, and the synergy site provides the mechanical strength to the bond.
journal_name
Biochemistryjournal_title
Biochemistryauthors
García AJ,Schwarzbauer JE,Boettiger Ddoi
10.1021/bi025752fsubject
Has Abstractpub_date
2002-07-23 00:00:00pages
9063-9issue
29eissn
0006-2960issn
1520-4995pii
bi025752fjournal_volume
41pub_type
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