Session TOE. There are 6 abstracts in this session.

Session: Biomolecular Structure & Function 1, time: 4:00-4:25
A contribution to integrated structural biology
Claudio Luchinat
CERM, Florence, Italy
Interpreting as real modest differences between X-ray and NMR structures is a risky attitude. We have proposed that the presence of real differences should be checked by a simultaneous refinement of the protein structure performed with both crystal and solution NMR data. If the two datasets are consistent with a single structural model, then the process provides a more accurate structure. To this purpose, the program REFMAC5 from CCP4 was modified (REFMAC-NMR) to allow the simultaneous use of X-ray crystallographic data and paramagnetic NMR data and/or diamagnetic residual dipolar couplings. Obvious advantages of this approach are that an independently solved solution structure is not needed, and also that the complementarity between X-ray and NMR data is fully exploited.

Session: Biomolecular Structure & Function 1, time: 4:25-4:50
Sponges and Fibrils:  Functional and Pathogenic Aggregates Examined by Solid-State NMR
Chad Rienstra
University of Illinois, Urbana,
In this talk I will present late-breaking results on my group's progress towards solution of high-resolution structures of the active solid-state aggregate of the gold standard antifungal drug amphotericin B and pathogenic fibrils of alpha-synuclein.  Particular attention will be paid to the completeness of SSNMR data with respect to structure models, approaches for leveraging high-dimensionality non-uniformly sampled data, structure determination from individual samples, and potential for automation of data analysis workflows.

Session: Biomolecular Structure & Function 1, time: 4:50-5:15
New Ways of Looking at Bacterial Cell Walls and Biofilms
Lynette Cegelski
Stanford University, Stanford, CA
The bacterial cell wall is essential to cell survival and is a major target of antibiotics. Beyond the cell surface, bacteria assemble macromolecular architectures during biofilm formation. Biofilms are implicated in serious infectious diseases and have emerged as a target for anti-infectives. Our research program is inspired by the challenge and importance of elucidating chemical structure and function in these complex biological systems and we strive to transform our discoveries into new therapeutic strategies. We have introduced new approaches using whole-cell and cell-wall solid-state NMR, integrated with biochemical analysis and rheology measurements, to reveal how the biological functions of cell walls and biofilms depend on their chemical composition and architecture. 

Session: Biomolecular Structure & Function 1, time: 5:15-5:30
Site-specific studies of nucleosome interactions by solid-state NMR
Shengqi Xiang; Ulric le Paige; Velten Horn; Klaartje Houben; Marc Baldus; Hugo Van Ingen
Utrecht University, Utrecht, Netherlands
Chromatin function depends on a dense network of interactions between nucleosomes and wide range of proteins. Given the great relevance of nucleosome-protein complexes, there is a demand for a diverse array of methods to study the molecular structure of these systems. We here present the first high-resolution study of nucleosome-protein interactions by state-of-the-art solid-state NMR (ssNMR).
Using sedimented nucleosomes, high-resolution spectra are obtained for both flexible histone tails and the non-mobile histone core, allowing the residue-specific study of the structure, dynamics and interactions of nucleosomes. Through co-sedimentation of a nucleosome-binding peptide, we demonstrate that protein binding sites on the nucleosome surface can be determined. We believe that this approach holds great promise in the study of nucleosome complexes.

Session: Biomolecular Structure & Function 1, time: 5:30-5:45
Structural investigation of the active site in a paramagnetic metalloprotein by fast magic angle spinning NMR
Andrea Bertarello1; Ladislav Benda1; Kevin Sanders1; Andrew Pell2; Vladimir Pelmenschikov4; Martin Kaupp4; Leonardo Gonnelli3; Isabella Felli3; Lyndon Emsley5; Roberta Pierattelli3; Guido Pintacuda1
1ENS Lyon-CNRS, Villeurbanne, France; 2Stockholm University, Stockholm, Sweden; 3CERM-University of Florence, Sesto Fiorentino, Italy; 4Technische Universit├Ąt Berlin, Berlin, Germany; 5EPFL, Lausanne, Switzerland
We show how the application of a set of NMR experiments at very fast (60-100 kHz) MAS, recently developed for the study of complex paramagnetic materials, can be adapted to the solid-state NMR analysis of paramagnetic metalloproteins. Working on a Co2+-enzyme, and complementing experiments with first-principles paramagnetic NMR calculations, we detect and assign signals from residues directly coordinating the metal centers in paramagnetic metalloenzymes. The obtained contact shifts are extremely sensitive to the fine details of the metal ion coordination, and constitute a powerful set of restraints for the determination of the structure of the metal site at sub-atomic resolution.

Session: Biomolecular Structure & Function 1, time: 5:45-6:00
Primary transfer step in the light-driven ion pump Bacteriorhodopsin: an irreversible U-turn revealed by DNP-enhanced MAS NMR
Qing Zhe Ni1; Thach Can1; Eugenio Daviso2; Marina Belenky2; Judith Herzfeld2; Robert Griffin1
1Massachusetts Institute of Technology, Cambridge, MA; 2Department of Chemistry, Brandeis University, Waltham, MA
Despite much attention, the path of the primary proton transfer in bacteriorhodopsin (bR) is still mysterious. Here we use DNP-enhanced MAS NMR to study the active site before the Schiff base (SB) deprotonates (L intermediate), after the SB deprotonates (Mo), and after the SB reprotonates (N). 13C-13C correlations show that D85 draws close to T89 before proton transfer and 15N(SB)-1H correlations reveal that the newly deprotonated SB is accepting from an alcohol. Concurrently, 15N-13C correlations between the SB and D85 show that helices C and G draw closer together and relax thereafter. Together, T89 serves to relay the SB proton to D85 and that creating this pathway involves rapprochement between the C and G helices.