Two- and three dimensional 1H NMR studies of a wheat phospholipid transfer protein: sequential resonance assignments and secondary structure

Two- and three-dimensional 1H NMR experiments have been used to sequentially assign nearly all proton resonances of the 90 residues of wheat phospholipid transfer protein. Only a few side-chain protons were not identified because of degeneracy or overlapping. The identification of spin systems and t...

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Bibliographic Details
Published in:Biochemistry (Easton) 1991-12, Vol.30 (49), p.11600-11608
Main Authors: Simorre, J.P. (Universite d'Orleans, Orleans, France), Caille, A, Marion, D, Ptak, M
Format: Article
Language:English
Subjects:
AGUA DEL SUELO
Amino Acid Sequence
Analytical, structural and metabolic biochemistry
Binding and carrier proteins
Biological and medical sciences
Carrier Proteins - chemistry
EAU DU SOL
Electron Spin Resonance Spectroscopy
ESPECTROMETRIA
ESPECTROSCOPIA RMN
FOSFOLIPIDOS
Fundamental and applied biological sciences. Psychology
GRAINE
Magnetic Resonance Spectroscopy
MEMBRANA
MEMBRANE
Membrane Proteins - chemistry
Molecular Sequence Data
PHOSPHATIDE
Phospholipid Transfer Proteins
Phospholipids - chemistry
Plant Proteins - chemistry
Protein Conformation
PROTEINAS VEGETALES
PROTEINE VEGETALE
Proteins
SEMILLA
SPECTROMETRIE
SPECTROSCOPIE RMN
Triticum - chemistry
TRITICUM AESTIVUM
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Summary:Two- and three-dimensional 1H NMR experiments have been used to sequentially assign nearly all proton resonances of the 90 residues of wheat phospholipid transfer protein. Only a few side-chain protons were not identified because of degeneracy or overlapping. The identification of spin systems and the sequential assignment were made at the same time by combining the data of the two- and three-dimensional experiments. The classical two-dimensional COSY, HOHAHA, and NOESY experiments benefit from both good resolution and high sensitivity, allowing the detection of long-range dipolar connectivities. The three-dimensional HOHAHA-NOESY experiment offers the advantage of a faster and unambiguous assignment. As a matter of fact, homonuclear three-dimensional NMR spectroscopy proved to be a very efficient method for resonance assignments of protein 1H NMR spectra which cannot be unraveled by 2D methods. An assignment strategy which overcomes most of the ambiguities has been proposed, in which each individual assignment toward the C-terminal end is supported by another in the opposite direction originating from a completely different part of the spectrum. Location of secondary structures of the phospholipid transfer protein was determined by using the method of analysis introduced here and was confirmed by 3J alpha NH coupling and NH exchange rates. Except for the C-terminal part, the polypeptide chain appears to be organized mainly as helical fragments connected by disulfide bridges. Further modeling will display the overall folding of the protein and should provide a better understanding of its interactions with lipids
ISSN:0006-2960
1520-4995