title: Studies on the solution conformation of human thioredoxin using heteronuclear 15N-1H nuclear magnetic resonance spectroscopy.
abstract: The solution conformation of uniformly labeled 15N human thioredoxin has been studied by two-dimensional heteronuclear 15N-1H nuclear magnetic resonance spectroscopy. Assignments of the 15N resonances of the protein are obtained in a sequential manner using heteronuclear multiple quantum coherence (HMQC), relayed HMQC-correlated (COSY), and relayed HMQC-nuclear Overhauser (NOESY) spectroscopy. Values of the 3JHN alpha splittings for 87 of the 105 residues of thioredoxin are extracted from a variant of the HMQC-COSY experiment, known as HMQC-J, and analyzed to give accurate 3JHN alpha coupling constants. In addition, long-range C alpha H(i)-15N(i + 1) scaler connectivities are identified by heteronuclear multiple bond correlation (HMBC) spectroscopy. The presence of these three-bond scaler connectivities in predominantly alpha-helical regions correlates well with the secondary structure determined previously from a qualitative analysis of homonuclear nuclear Overhauser data [Forman-Kay, J. D., Clore, G. M., Driscoll, P.C., Wingfield, P. T., Richards, F. M., & Gronenborn, A. M. (1989) Biochemistry 28, 7088-7097], suggesting that this technique may provide additional information for secondary structure determination a priori. The accuracy with which 3JHN alpha coupling constants can be obtained from the HMQC-J experiment permits a more precise delineation of the beginnings and ends of secondary structural elements of human thioredoxin and of irregularities in these elements.
authors.affiliation: Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: J E
authors.initials: JE
authors.lastname: Forman-Kay
authors.forename: A M
authors.initials: AM
authors.lastname: Gronenborn
authors.forename: L E
authors.initials: LE
authors.lastname: Kay
authors.forename: P T
authors.initials: PT
authors.lastname: Wingfield
authors.forename: G M
authors.initials: GM
authors.lastname: Clore
mesh_terms: D001426:Bacterial Proteins; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D009682:Magnetic Resonance Spectroscopy; D011487:Protein Conformation; D012996:Solutions; D013879:Thioredoxins
chemical_list: D001426:Bacterial Proteins; D012996:Solutions; D013879:Thioredoxins