Miocene waterfowl and other birds from central Otago, New Zealand

Copyright © The Natural History Museum 2007Abundant fossil bird bones from the lower Bannockburn Formation, Manuherikia Group, an Early-Middle Miocene lacustrine deposit, 16–19 Ma, from Otago in New Zealand, reveal the “St Bathans Fauna” (new name), a first Tertiary avifauna of land and freshwater birds from New Zealand. At least 23 species of birds are represented by bones, and probable moa, Aves: Dinornithiformes, by eggshell. Anatids dominate the fauna with four genera and five species described as new: a sixth and largest anatid species is represented by just one bone. This is the most diverse Early-Middle Miocene duck fauna known worldwide. Among ducks, two species of dendrochenines are most numerous in the fauna, but a tadornine is common as well. A diving petrel (Pelecanoididae: Pelecanoides) is described, so extending the geological range of this genus worldwide from the Pliocene to the Middle Miocene, at least. The remaining 16 taxa are left undescribed but include: a large species of gull (Laridae); two small waders (Charadriiformes, genus indet.), the size of Charadrius bicinctus and Calidris ruficollis, respectively; a gruiform represented by one specimen similar to Aptornis; abundant rail (Rallidae) bones, including a common flightless rail and a rarer slightly larger taxon, about the size of Gallirallus philippensis; an ?eagle (Accipitridae); a pigeon (Columbidae); three parrots (Psittacidae); an owlet nightjar (Aegothelidae: Aegotheles sp.); a swiftlet (Apodidae: Collocalia sp.); and three passerine taxa, of which the largest is a member of the Cracticidae. The absence of some waterbirds, such as anserines (including swans), grebes (Podicipedidae) and shags (Phalacrocoracidae), among the abundant bones, indicates their probable absence from New Zealand in the Early-Middle Miocene.T. H. Worthy, A. J. D. Tennyson, C. Jones, J. A. McNamara and B. J. Dougla

Minimum Free Energy Path of Ligand-Induced Transition in Adenylate Kinase

Large-scale conformational changes in proteins involve barrier-crossing transitions on the complex free energy surfaces of high-dimensional space. Such rare events cannot be efficiently captured by conventional molecular dynamics simulations. Here we show that, by combining the on-the-fly string method and the multi-state Bennett acceptance ratio (MBAR) method, the free energy profile of a conformational transition pathway in Escherichia coli adenylate kinase can be characterized in a high-dimensional space. The minimum free energy paths of the conformational transitions in adenylate kinase were explored by the on-the-fly string method in 20-dimensional space spanned by the 20 largest-amplitude principal modes, and the free energy and various kinds of average physical quantities along the pathways were successfully evaluated by the MBAR method. The influence of ligand binding on the pathways was characterized in terms of rigid-body motions of the lid-shaped ATP-binding domain (LID) and the AMP-binding (AMPbd) domains. It was found that the LID domain was able to partially close without the ligand, while the closure of the AMPbd domain required the ligand binding. The transition state ensemble of the ligand bound form was identified as those structures characterized by highly specific binding of the ligand to the AMPbd domain, and was validated by unrestrained MD simulations. It was also found that complete closure of the LID domain required the dehydration of solvents around the P-loop. These findings suggest that the interplay of the two different types of domain motion is an essential feature in the conformational transition of the enzyme

Fast and Accurate Resonance Assignment of Small-to-Large Proteins by Combining Automated and Manual Approaches

The process of resonance assignment is fundamental to most NMR studies of protein structure and dynamics. Unfortunately, the manual assignment of residues is tedious and time-consuming, and can represent a significant bottleneck for further characterization. Furthermore, while automated approaches have been developed, they are often limited in their accuracy, particularly for larger proteins. Here, we address this by introducing the software COMPASS, which, by combining automated resonance assignment with manual intervention, is able to achieve accuracy approaching that from manual assignments at greatly accelerated speeds. Moreover, by including the option to compensate for isotope shift effects in deuterated proteins, COMPASS is far more accurate for larger proteins than existing automated methods. COMPASS is an open-source project licensed under GNU General Public License and is available for download from http://www.liu.se/forskning/foass/tidigare-foass/patrik-lundstrom/software?l=en. Source code and binaries for Linux, Mac OS X and Microsoft Windows are available.Funding Agencies|Swedish Research Council [Dnr. 2012-5136]</p

Application of Methyl-TROSY to a Large Paramagnetic Membrane Protein Without Perdeuteration: \u3csup\u3e13\u3c/sup\u3eC-MMTS-labeled NADPH-cytochrome P450 Oxidoreductase

NMR spectroscopy of membrane proteins involved in electron transport is difficult due to the presence of both the lipids and paramagnetic centers. Here we report the solution NMR study of the NADPH-cytochrome P450 oxidoreductase (POR) in its reduced and oxidized states. We interrogate POR, first, in its truncated soluble form (70 kDa), which is followed by experiments with the full-length protein incorporated in a lipid nanodisc (240 kDa). To overcome paramagnetic relaxation in the reduced state of POR as well as the signal broadening due to its high molecular weight, we utilized the methyl-TROSY approach. Extrinsic 13C-methyl groups were introduced by modifying the engineered surface-exposed cysteines with methyl-methanethiosulfonate. Chemical shift dispersion of the resonances from different sites in POR was sufficient to monitor differential effects of the reduction–oxidation process and conformation changes in the POR structure related to its function. Despite the high molecular weight of the POR-nanodisc complex, the surface-localized 13C-methyl probes were sufficiently mobile to allow for signal detection at 600 MHz without perdeuteration. This work demonstrates a potential of the solution methyl-TROSY in analysis of structure, dynamics, and function of POR, which may also be applicable to similar paramagnetic and flexible membrane proteins