Amino acid type determination in the sequential assignment procedure of uniformly 13C/15N-enriched proteins.
Experiments and procedures are described that greatly alleviate the sequential assignment process of uniformly 13C/15N-enriched proteins by determining the type of amino acid from experiments that correlate side chain with backbone amide resonances. A recently proposed 3D NMR experiment, CBCA(CO)NH, correlates C alpha and C beta resonances to the backbone amide 1H and 15N resonances of the next residue (Grzesiek, S. and Bax, A. (1992) J. Am. Chem. Soc., 114, 6291-6293). An extension of this experiment is described which correlates the proton H beta and H alpha resonances to the amide 1H and 15N resonances of the next amino acid, and a detailed product operator description is given. A simple 2D-edited constant-time HSQC experiment is described which rapidly identifies H beta and C beta resonances of aromatic or Asn/Asp residues. The extent to which combined knowledge of the C alpha and C beta chemical shift values determines the amino acid type is investigated, and it is demonstrated that the combined C alpha and C beta chemical shifts of three or four adjacent residues usually are sufficient for defining a unique position in the protein sequence.
Grzesiek
S
S
Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.
Bax
A
A
D000465:Algorithms; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D002147:Calmodulin / Q000737:chemistry; D007371:Interferon-gamma / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009219:Myosin-Light-Chain Kinase / Q000737:chemistry; D010446:Peptide Fragments / Q000737:chemistry; D011485:Protein Binding; D012984:Software
D000596:Amino Acids; D002147:Calmodulin; D010446:Peptide Fragments; D007371:Interferon-gamma; D009219:Myosin-Light-Chain Kinase