TROSY experiment for refinement of backbone psi and phi by simultaneous measurements of cross-correlated relaxation rates and 3,4J(H alpha HN) coupling constants.
The TROSY principle has been introduced into a HNCA experiment, which is designed for measurements of the intraresidual and sequential H(alpha)-C(alpha)/H(N)-N dipole/dipole and H(alpha)-C(alpha)/N dipole/CSA cross-correlated relaxation rates. In addition, the new experiment provides values of the (3,4)J(H alpha HN) coupling constants measured in an E.COSY manner. The conformational restraints for the psi and phi angles are obtained through the use of the cross-correlated relaxation rates together with the Karplus-type dependencies of the coupling constants. Improved signal-to-noise is achieved through preservation of all coherence transfer pathways and application of the TROSY principle. The application of the [(15)N,(13)C]-DQ/ZQ-[(15)N,(1)H]-TROSY-E.COSY experiment to the 16 kDa apo-form of the E. coli Heme Chaperon protein CcmE is described. Overall good agreement is achieved between psi and phi angles measured with the new experiment and the average values determined from an ensemble of 20 NMR conformers.
Vögeli
Beat
B
Laboratorium für Physikalische Chemie, Swiss Federal Institute of Technology, ETH-Hönggerberg, CH-8093 Zürich, Switzerland.
Pervushin
Konstantin
K
D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry*; D002247:Carbon Isotopes; D029968:Escherichia coli Proteins / Q000737:chemistry; D006420:Hemeproteins / Q000737:chemistry*; D008280:Magnetics; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D017433:Protein Structure, Secondary
D001425:Bacterial Outer Membrane Proteins; D002247:Carbon Isotopes; C115384:CcmE protein, E coli; D029968:Escherichia coli Proteins; D006420:Hemeproteins; D009587:Nitrogen Isotopes