Abstract
Crystal structures of classical cadherins have revealed two dimeric configurations. In the first, N-terminal β-strands of EC1 domains 'swap' between partner molecules. The second configuration (the 'X dimer'), also observed for T-cadherin, is mediated by residues near the EC1-EC2 calcium binding sites, and N-terminal β-strands of partner EC1 domains, though held adjacent, do not swap. Here we show that strand-swapping mutants of type I and II classical cadherins form X dimers. Mutant cadherins impaired for X-dimer formation show no binding in short–time frame surface plasmon resonance assays, but in long–time frame experiments, they have homophilic binding affinities close to that of wild type. Further experiments show that exchange between monomers and dimers is slowed in these mutants. These results reconcile apparently disparate results from prior structural studies and suggest that X dimers are binding intermediates that facilitate the formation of strand-swapped dimers.
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Acknowledgements
This work was supported in part by US National Institutes of Health grants R01 GM062270 (L.S.), R01 GM30518 (B.H.) and U54 CA121852 (B.H. and L.S.) and by US National Science Foundation grant MCB-0416708 (B.H.). B.H. is an investigator of the Howard Hughes Medical Institute. X-ray data were acquired at the X4A and X4C beamlines of the National Synchrotron Light Source, Brookhaven National Laboratory; the X4 beamlines are operated by the New York Structural Biology Center.
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O.J.H., X.J., F.B. and K.J.C. determined and refined all the crystal structures; F.B., O.J.H. and J.B. produced all the wild-type and mutant proteins; P.S.K. performed and analyzed the SPR experiments; J.V. performed bioinformatic analysis; G.A. performed the AUC experiments; S.R.P. performed the cell-aggregation and immunofluorescence studies; O.J.H., B.H. and L.S. designed experiments, analyzed data and wrote the manuscript.
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Harrison, O., Bahna, F., Katsamba, P. et al. Two-step adhesive binding by classical cadherins. Nat Struct Mol Biol 17, 348–357 (2010). https://doi.org/10.1038/nsmb.1784
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DOI: https://doi.org/10.1038/nsmb.1784
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