Deposits of clastic carbonate-dominated (calciclastic) sedimentary slope systems in the rock record have been identified mostly as linearly-consistent carbonate apron deposits, even though most ancient clastic carbonate slope deposits fit the submarine fan systems better. Calciclastic submarine fans are consequently rarely described and are poorly understood. Subsequently, very little is known especially in mud-dominated calciclastic submarine fan systems. Presented in this study are a sedimentological core and petrographic characterisation of samples from eleven boreholes from the Lower Carboniferous of Bowland Basin (Northwest England) that reveals a >250 m thick calciturbidite complex deposited in a calciclastic submarine fan setting. Seven facies are recognised from core and thin section characterisation and are grouped into three carbonate turbidite sequences. They include: 1) Calciturbidites, comprising mostly of highto low-density, wavy-laminated bioclast-rich facies; 2) low-density densite mudstones which are characterised by planar laminated and unlaminated muddominated facies; and 3) Calcidebrites which are muddy or hyper-concentrated debrisflow deposits occurring as poorly-sorted, chaotic, mud-supported floatstones. These

Phd by thesis

5.1. Binding Sites of the Aminoglycosides 1185 5.2. Importance of Electrostatic Interactions 1185 5.3. Nonionic Interactions 1185 5.4. Pseudo-Base Pair Interactions 1186 5.5. Water-Mediated Contacts 1186 5.6. Shape Complementarity and Conformational Adaptation 1186 6. Assays for Evaluating RNA Binding 1186 6.1. Methods Utilizing Fluorescently Labeled RNA 1186 6.2. NMR-Based Methods 1187 6.3. Electrospray Ionization Mass Spectrometry-Based Methods 1187

Targeting RNA with small molecules

Enzyme catalysis is an inherently dynamic process. Binding and release of ligands is often accompanied by conformational changes, both subtle and dramatic (reviewed more extensively in refs 1 -4), and these conformational changes may be rate-limiting in the overall reaction scheme, for example in dihydrofolate reductase (DHFR), 5 triosephosphate isomerase (TIM), 6,7 and protein kinase A. 8 The catalytic step itself, by its very nature, is dynamic. Atomic fluctuations along the reaction coordinate lead to bonds being broken and new ones being formed. Binding of substrate can lead to the repositioning of catalytic groups, effectively bridging the dynamic processes of substrate binding and catalysis. 2,9

An NMR perspective on enzyme dynamics.

1.1. Why Study Membranes and Membrane Proteins?
Biological membranes and membrane proteins, responsible for numerous exciting biological processes, present one of the paramount challenges in biophysics today. Membranes are present in great number and variety in all organisms. They form the boundary between the inside and outside for any bacterium or cell, and they delimit the host of organelles that make up their inner subunits. Each biological membrane is made up of dozens of different types of lipids and sterols, and any particular type of membrane has a characteristic content of these different constituents. As a very basic example, we mention that prokaryotic membranes contain a notable component of negatively charged lipids but almost no cholesterol, while eukaryotic membranes are mostly zwitterionic but have a significant amount of cholesterol. Since the driving biophysical principles of membrane formation are very simple—they lie in the amphipathic properties of any lipid molecule—a single lipid type is sufficient to form membrane-like bilayers in an aqueous environment. Such model membranes are used extensively to study biophysical properties that are representative for most membrane systems. A particularly interesting effect is observed when detergent molecules are added to lipid bilayer samples: the detergents solubilize the bilayers, and in certain regimes so-called bilayered mixed micelles or “bicelles” are formed. In the simplest case, they can be described as microscopic disks where a bilayer patch is encircled by a “rim” of detergent molecules. Bicelles represent a new instance of lipid morphology and are extensively applicable to structural studies of lipid membranes and protein structure.1

Membranes delimit any cell and all of its compartments. They form natural borders for metabolic substances and signaling molecules. Membrane proteins are the porters and gatekeepers that make sure that only proper molecules or signals make it across the membrane. Since membrane proteins perform numerous key functions in cell metabolism and signaling, they contribute over 30% of the genes in typical eukaryotic genomes,2 and they form the targets for over 50% of drugs in use today.3 The number of elucidated structures of membrane proteins has grown exponentially after the first structure was published in 1985, thus equaling the rate at which structure determination of soluble proteins emerged early on.4 Still, the number of available high-resolution structures of membrane proteins is limited. There are Internet sites that keep track of newly published structures of membrane proteins. The crystallography-oriented Web site of Dr. Stephen White [http://blanco.biomol.uci.edu/mpstruc] has recently been joined by another site maintained by Dr. Dror Warschawski that is dedicated to structures of membrane proteins elucidated by nuclear magnetic resonance (NMR) spectroscopy [www.drorlist.com/nmr/MPNMR.html]. Another equally important site of Dr. Hartmut Michel [www.mpibp-frankfurt.mpg.de/michel/public/memprotstruct.html] with an emphasis on crystallization conditions is no longer updated, but states that access is still enabled.

In this review article, we aim to give a general overview of lipid bicelles as employed in the study of protein structure. Recent advances in the field of protein structural biology that have been made possible by exploiting the unique properties of lipid bicelles, in both solution and solid-state NMR spectroscopy, will be discussed. During the last five years, review contributions have presented bicelles either within the far more general context of reconstitution media for solution NMR studies (see section 1.4) or have focused on macroscopically aligned bicelles as used for solid-state NMR studies.5,6 One very recent contribution has tackled the formidable task of reviewing all membrane mimetics employed in both solution and solid-state NMR studies.7 As mentioned above, we will limit the contents of this review article to applications of lipid bicelles, but will cover both the isotropic and the aligned bicelles as used in NMR studies. Some parts of this article can be viewed as an update on the review articles of Opella and Marassi,8 Marcotte and Auger,9 and Prosser et al.10 In addition, some of our own recent research involving bicelles is presented in detail.

The magic of bicelles lights up membrane protein structure.

N(6)-Methyladenosine Modification in a Long Noncoding RNA Hairpin Predisposes Its Conformation to Protein Binding

Proper functioning of RNAs requires the formation of complex three-dimensional structures combined with the ability to rapidly interconvert between multiple functional states. This review covers recent advances in isotope-labeling strategies and NMR experimental approaches that have promise for facilitating solution structure determinations and dynamics studies of biologically active RNAs. Improved methods for the production of isotopically labeled RNAs combined with new multidimensional heteronuclear NMR experiments make it possible to dramatically reduce spectral crowding and simplify resonance assignments for RNAs. Several novel applications of experiments that directly detect hydrogen-bonding interactions are discussed. These studies demonstrate how NMR spectroscopy can be used to distinguish between possible secondary structures and identify mechanisms of ligand binding in RNAs. A variety of recently developed methods for measuring base and sugar residual dipolar couplings are described. NMR residual dipolar coupling techniques provide valuable data for determining the long-range structure and orientation of helical regions in RNAs. A number of studies are also presented where residual dipolar coupling constraints are used to determine the global structure and dynamics of RNAs. NMR relaxation data can be used to probe the dynamics of macromolecules in solution. The power dependence of transverse rotating-frame relaxation rates was used here to study dynamics in the minimal hammerhead ribozyme. Improved methods for isotopically labeling RNAs combined with new types of structural data obtained from a growing repertoire of NMR experiments are facilitating structural and dynamic studies of larger RNAs.

NMR Methods for Studying the Structure and Dynamics of RNA

Residual dipolar couplings (RDCs) complement standard NOE distance and J-coupling torsion angle data to improve the local and global structure of biomolecules in solution. One powerful application of RDCs is for domain orientation studies, which are especially valuable for structural studies of nucleic acids, where the local structure of a double helix is readily modeled and the orientations of the helical domains can then be determined from RDC data. However, RDCs obtained from only one alignment media generally result in degenerate solutions for the orientation of multiple domains. In protein systems, different alignment media are typically used to eliminate this orientational degeneracy, where the combination of RDCs from two (or more) independent alignment tensors can be used to overcome this degeneracy. It is demonstrated here for native E. coli tRNA(Val) that many of the commonly used liquid crystalline alignment media result in very similar alignment tensors, which do not eliminate the 4-fold degeneracy for orienting the two helical domains in tRNA. The intrinsic magnetic susceptibility anisotropy (MSA) of the nucleobases in tRNA(Val) was also used to obtain RDCs for magnetic alignment at 800 and 900 MHz. While these RDCs yield a different alignment tensor, the specific orientation of this tensor combined with the high rhombicity for the tensors in the liquid crystalline media only eliminates two of the four degenerate orientations for tRNA(Val). Simulations are used to show that, in optimal cases, the combination of RDCs obtained from liquid crystalline medium and MSA-induced alignment can be used to obtain a unique orientation for the two helical domains in tRNA(Val).

Darin J. Brown

Papers

NMR Methods for Studying the Structure and Dynamics of RNA

Comparison of alignment tensors generated for native tRNA(Val) using magnetic fields and liquid crystalline media.