1H and 15N resonance assignments and secondary structure of cellular retinoic acid-binding protein with and without bound ligand

Jose Rizo-Rey, Zhi-Ping Liu, Lila M. Gierasch

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Sequence-specific assignments for the 1H and 15N backbone resonances of cellular retinoic acid-binding protein (CRABP), with and without the bound ligand, have been obtained. Most of the side-chain resonances of both apo- and holo-CRABP have also been assigned. The assignments have been obtained using two-dimensional homonuclear and heteronuclear NMR data, and three-dimensional 1H-15N TOCSY-HMQC and NOESY-HMQC experiments. The secondary structure, deduced from nuclear Overhauser effects, amide H/D exchange rates and Hα chemical shifts, is analogous in both forms of the protein and is completely consistent with a model of CRABP that had been constructed by homology with the crystal structure of myelin P2 protein [Zhang et al. (1992) Protein Struct. Funct. Genet., 13, 87-99]. This model comprises two five-stranded β-sheets that form a sandwich or β-clam structure, and a short N-terminal helix-turn-helix motif that closes the binding cavity between the two sheets. Comparison of the data obtained for apo- and holo-CRABP indicates that a region around the C-terminus of the second helix is much more flexible in the apo-protein. Our data provide experimental evidence for the hypothesis that the ligand-binding mechanism of CRABP, and of other homologous proteins that bind hydrophobic ligands in the cytoplasm, involves opening of a portal to allow entry of the ligand into the cavity.

Original languageEnglish (US)
Pages (from-to)741-760
Number of pages20
JournalJournal of Biomolecular NMR
Issue number6
StatePublished - Nov 1 1994



  • Cellular retinoic acid binding protein
  • Multidimensional NMR
  • Resonance assignment
  • Secondary structure
  • Selective labeling

ASJC Scopus subject areas

  • Biochemistry
  • Spectroscopy

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