1H, 13C and 15N assignments and chemical shift-derived secondary structure of intestinal fatty acid-binding protein.
Sequence-specific 1H, 13C and 15N resonance assignments have been established for rat intestinal fatty acid-binding protein complexed with palmitate (15.4 kDa) at pH 7.2 and 37 degrees C. The resonance assignment strategy involved the concerted use of seven 3D triple-resonance experiments (CC-TOCSY, HCCH-TOCSY, HNCO, HNCA, 15N-TOCSY-HMQC, HCACO and HCA(CO)N). A central feature of this strategy was the concurrent assignment of both backbone and side-chain aliphatic atoms, which was critical for overcoming ambiguities in the assignment process. The CC-TOCSY experiment provided the unambiguous links between the side-chain spin systems observed in HCCH-TOCSY and the backbone correlations observed in the other experiments. Assignments were established for 124 of the 131 residues, although 6 of the 124 had missing amide 1H resonances, presumably due to rapid exchange with solvent under these experimental conditions. The assignment database was used to determine the solution secondary structure of the complex, based on chemical shift indices for the 1H alpha, 13C alpha, 13C beta and 13CO atoms. Overall, the secondary structure agreed well with that determined by X-ray crystallography [Sacchettini et al. (1989) J. Mol. Biol., 208, 327-339], although minor differences were observed at the edges of secondary structure elements.
Hodsdon
M E
ME
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO 63110-1010, USA.
Toner
J J
JJ
Cistola
D P
DP
D000595:Amino Acid Sequence; D000818:Animals; D002247:Carbon Isotopes; D002352:Carrier Proteins / Q000737:chemistry*; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D006859:Hydrogen; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D018996:Myelin P2 Protein / Q000737:chemistry*; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D009587:Nitrogen Isotopes; D017433:Protein Structure, Secondary; D051381:Rats
D002247:Carbon Isotopes; D002352:Carrier Proteins; C490174:Fabp7 protein, rat; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D018996:Myelin P2 Protein; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D009587:Nitrogen Isotopes; D006859:Hydrogen