NMR structure and dynamics of human ephrin-B2 ectodomain: the functionally critical C-D and G-H loops are highly dynamic in solution.
Eph receptors and ephrins constitute the largest family of receptor tyrosine kinases with 15 individual receptors and nine ligands. Its ectodomains represent attractive targets not only for understanding fundamental mechanisms underlying axon guidance, cell migration, segmentation, tumorigenesis, and bone remodeling, but also for drug screening/design to treat cancers, bone diseases and viral infection. So far no NMR study on the ephrin ectodomains is available and as such their properties in solution still remain unknown. In this study, we presented the first NMR structure and dynamics of the human ephrin-B2 ectodomain as well as its interaction with the receptor EphB2. Strikingly, the NMR study reveals a picture different from those previously obtained by X-ray crystallography. Although in solution it still adopts the same Greek key fold, with the central beta-barrel ( approximately 30% of the molecule) highly similar to that in crystal structures, the other regions are highly dynamic and accessible to the bulk solvent. In particular, the functionally critical C-D and G-H loops of the ephrin-B2 ectodomain are highly flexible as reflected by several NMR probes including hydrogen exchange and (15)N backbone relaxation data. Nevertheless, as revealed by ITC and NMR, the ephrin-B2 ectodomain binds to EphB2 with a K(d) of 22.3 nM to form a tight complex in which the tip of the C-D loop and the C-terminus still remain largely flexible. The present results may bear critical implications in understanding the molecular details as well as designing antagonists of therapeutic interest for Eph-ephrin interactions.
Ran
Xiaoyuan
X
Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
Qin
Haina
H
Liu
Jingxian
J
Fan
Jing-Song
JS
Shi
Jiahai
J
Song
Jianxing
J
D002151:Calorimetry; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D003903:Deuterium; D036388:Ephrin-B2 / Q000737:chemistry*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D009584:Nitrogen; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D036183:Receptor, EphB2 / Q000737:chemistry; D012996:Solutions
D036388:Ephrin-B2; D012996:Solutions; D003903:Deuterium; D036183:Receptor, EphB2; D009584:Nitrogen