Code: ThOE	Time Slot/Poster Number: 4:00-4:25	Session: Dynamics & Computation
Recent Advances of MAS Solid-state NMR using Perdeuterated Peptides and Proteins
Veniamin Chevelkov1; Vipin Agarwal1; Rasmus Linser1; Andi Mainz1; Juan-Miguel Lopez del Amo1; Muralidhar Dasari1; Uwe Fink1; Yi Xue2; Nikolai R. Skrynnikov2; Bernd Reif1
1FMP Berlin, Berlin, Germany; 2Purdue University, West Lafayette, USA
Abstract
MAS solid-state NMR in isotopically enriched materials is limited by resolution and sensitivity. We employ perdeuteration and back-substitution of exchangeable protons in peptides and proteins in order to reduce 1H, 1H dipolar interactions. This labeling scheme enables also the observation of side chain exchangeable protons which are easily assigned making use of dipolar couplings for magnetization transfer. Similarly, high sensitivitiy and high resolution spectra are obtained for methyl containing side chains by either incorporating specifically protonated precursors or by making use of the residual protons in otherwise perdeuterated proteins. We present relaxation experiments to characterize backbone and side chain dynamics in the solid-state.

Code: ThOE	Time Slot/Poster Number: 4:25-4:40	Session: Dynamics & Computation
Structure-Independent Determination of Distribution Radius for Disordered Group in Macromolecule by NMR: Use of Order Parameters for Long Variable-Length Vectors
Junji Iwahara1; G. Marius Clore2
1University of Texas Medical Branch, Galveston, TX; 2Laboratory of Chemical Physics, NIDDK, NIH, Bethesda, MD
Abstract
Distribution radii of disordered groups in macromolecules are very difficult to determine by any experiments. Although NMR order parameters for bond vectors (e.g. N-H and C-H) are commonly used to analyze conformational dynamics of macromolecules, these parameters are totally insensitive to translational motions. Here we demonstrate a novel method for structure-independent determination of distribution radii of disorder groups in macromolecules. The key concept is the use of the order parameters for long variable-length vectors between an atom in the disordered group and another atom at a rigid part of the macromolecule. We have developed the formalism to obtain the distribution radii from this kind of order parameters, and applied it to vectors for NMR paramagnetic relaxation enhancement.

Code: ThOE	Time Slot/Poster Number: 4:40-5:05	Session: Dynamics & Computation
NMR Relaxation Experiments for Proteins at High-Magnetic Field Strength
Rieko Ishima
University of Pittsburgh, Pittsburgh, PA
Abstract
High-field NMR provides better signal separation that is suitable for analysis of large proteins, and field dependent experiments are useful to determine dynamics parameters accurately. In contrast, at high magnetic field strength, spin-lock, CPMG R2, or its relaxation dispersion version requires high radio-frequency power. Due to increase in the resonance frequency as increase in the magnetic field strength, actual signal-to-noise ratio may decrease by increase in the transverse relaxation rate or the decrease in the longitudinal relaxation rate. Thus, we need to optimize the relaxation experiments and the analysis methods. We describe our recent modification of NMR relaxation data analysis and our simulations results to clarify the effects of different magnetic field strengths.

Code: ThOE	Time Slot/Poster Number: 5:05-5:20	Session: Dynamics & Computation
Measuring kinetics of conformational sampling in ubiquitin: Implications for protein-protein recognition
Donghan Lee1; Michael Funk1; Rudolf Gulich2; Dalia Egger2; Korvin Walter1; Bryn Fenwick3; Karin Giller1; Fabio Pichierri4; Bert de Groot5; Oliver Lange5; Helmut Grubmüller5; Xavier Salvatella3; Alois Loidl2; Reiner Kree6; Stefan Becker1; Nils Lakomek1; Peter Lunkenheimer2; Christian Griesinger1
1Max Planck Institute for Biophysical Chemistry, Göttingen, Germany; 2University of Augsburg, Experimental Physics V, Augsburg, Germany; 3Institute for Research in Biomedicine, Barcelona, Spain; 4G-COE Laboratory, Dept. of Applied Chemistry, Sendai, Japan; 5Dept. for Theoretical Biophysics, MPIBPC, Gottingen, Germany; 6Institute for Theoretical Physics, U Gottingen, Gottingen, Germany
Abstract
RDC-enhanced NMR spectroscopy has recently detected protein dynamics in a hidden time window (supra-τc) to play a major role for protein-protein recognition in a conformational selection scenario. We determine the lifetime of the different conformations in the solution ensemble to be about 1 μs at 309 K and approximately 50 μs at 265 K. This result is obtained by dielectric relaxation (DR) spectroscopy via a newly discovered mechanism of coupling conformational variation to the ion mobility. By considering RDC-derived ensembles and the time scale of inter-conversion between the different conformations as measured by DR spectroscopy, we can predict NMR relaxation dispersion data of ubiquitin in super-cooled water, thus providing evidence for the 1 μs supra-τc motion with atomic resolution.

Code: ThOE	Time Slot/Poster Number: 5:20-5:45	Session: Dynamics & Computation
Joint Analysis of Conformational Dynamics in Ribonuclease H using NMR Spectroscopy and Molecular Dynamics Simulations
Arthur G Palmer
Columbia University, New York, NY
Abstract
Homologous mesophilic (E. coli) and thermophilic (T. thermophilus) ribonuclease H (RNase H) enzymes illustrate how changes in protein sequence and structure that affect conformational dynamic processes can be monitored and characterized by joint analysis of NMR spectroscopy and molecular dynamics simulations. Specific intramolecular interactions that modulate backbone and sidechain dynamical properties of RNase H proteins have been identified using MD simulations and subsequently confirmed by NMR spin relaxation measurements. These studies emphasize the importance of hydrogen bonds and local steric interactions in restricting conformational fluctuations, and the absence of such interactions in allowing conformational adaptation to substrate binding.

Code: ThOE	Time Slot/Poster Number: 5:45-6:00	Session: Dynamics & Computation
A Simple Analytic Formalism Accounts for 13C and 15N T1 Relaxation of Solid Proteins and Peptides Under MAS
Elizabeth Fry1; Van Phan1; Andrew Byrd2; Kurt Zilm1
1Yale University, New Haven, CT; 2Structural Biophysics Laboratory, NCI, Frederick, MD
Abstract
An analytic formalism has been developed for extracting dynamics parameters for proteins in the solid state from 15N and 13C T1 measurements under MAS. The method has been validated by studying methyl group 13C relaxation where the rotational correlation time is well known to closely follow an Arrhenius temperature dependence. In application to ubiquitin, the dispersion in amide 15N T1s is found to be a result of the variability of 15N-1H librations. The average correlation time tau is found to be 30 ps, with a range of 10 ps to 60 ps observed.