Cobalamin (Cbl, vitamin B12) is a bacterial organic compound and an essential coenzyme in mammals, which take it up from the diet. This occurs by the combined action of the gastric intrinsic factor (IF) and the ileal endocytic cubam receptor formed by the 460-kilodalton (kDa) protein cubilin and the 45-kDa transmembrane protein amnionless1,2. Loss of function of any of these proteins ultimately leads to Cbl deficiency in man3,4. Here we present the crystal structure of the complex between IF–Cbl and the cubilin IF–Cbl-binding-region (CUB5–8)5 determined at 3.3 Å resolution. The structure provides insight into how several CUB (for ‘complement C1r/C1s, Uegf, Bmp1’) domains collectively function as modular ligand-binding regions, and how two distant CUB domains embrace the Cbl molecule by binding the two IF domains in a Ca2+-dependent manner. This dual-point model provides a probable explanation of how Cbl indirectly induces ligand–receptor coupling. Finally, the comparison of Ca2+-binding CUB domains and the low-density lipoprotein (LDL) receptor-type A modules suggests that the electrostatic pairing of a basic ligand arginine/lysine residue with Ca2+-coordinating acidic aspartates/glutamates is a common theme of Ca2+-dependent ligand–receptor interactions.
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We are grateful to G. Biller, G. Ratz and G. Hartvigsen for technical assistance, the staff at MAX-lab and SLS beamlines for help with data collection, and S. Fedosov for advice on expression. The study was essentially supported by a grant to S.K.M. from the Lundbeck foundation. G.R.A. was supported by the Danish Science Research Council and a Hallas-Møller stipend from the Novo-Nordisk foundation. M.M. was supported by the Novo Nordisk Foundation.
Author Contributions C.B.F.A.: cloning, expression, purification, crystallization, data collection, structure determination and analysis, manuscript preparation; M.M.: cloning, expression, purification, size-exclusion chromatographic analyses, manuscript preparation; T.S.: cloning, mutagenesis, expression, size-exclusion chromatographic analyses, cross-linkage experiments; G.R.A. and S.K.M.: study design and manuscript preparation.
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Andersen, C., Madsen, M., Storm, T. et al. Structural basis for receptor recognition of vitamin-B12–intrinsic factor complexes. Nature 464, 445–448 (2010). https://doi.org/10.1038/nature08874
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