研究者詳細 - TAMA Florence

論文 - TAMA Florence

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ELASTIC IMAGE REGISTRATION TO FULLY EXPLORE MACROMOLECULAR DYNAMICS BY ELECTRON MICROSCOPY 査読有り

Qiyu Jin, Carlos Oscar Sanchez Sorzano, Isabelle Callebaut, Florence Tama, Slavica Jonic

2014 IEEE INTERNATIONAL CONFERENCE ON IMAGE PROCESSING (ICIP) 頁： 2075 - 2079 2014年

記述言語：英語掲載種別：研究論文（国際会議プロシーディングス）出版者・発行元：IEEE

Structural changes are critical for biological functions of proteins and describing conformational changes in large macromolecular complexes is a major challenge. We have recently developed a hybrid method (HEMNMA) combining transmission electron microscopy (EM), normal mode analysis (NMA), and image analysis to study macromolecular dynamics. NMA is traditionally used to study macromolecular motions while HEMNMA provides insight into actual conformational changes seen by EM. HEMNMA uses normal modes to elastically align EM images with a reference structure in order to determine the conformations present in images and evaluate their pertinence. In this paper, we show how HEMNMA can be used with an atomic-resolution reference structure, using as an example the study of the conformational dynamics of Tomato Bushy Stunt Virus.

Allosteric Regulation of DNA Cleavage and Sequence-Specificity through Run-On Oligomerization 査読有り

Dmitry Lyumkis, Heather Talley, Andrew Stewart, Santosh Shah, Chad K. Park, Florence Tama, Clinton S. Potter, Bridget Carragher, Nancy C. Horton

STRUCTURE 21 巻 ( 10 ) 頁： 1848 - 1858 2013年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：CELL PRESS

SgrAl is a sequence specific DNA endonuclease that functions through an unusual enzymatic mechanism that is allosterically activated 200- to 500-fold by effector DNA, with a concomitant expansion of its DNA sequence specificity. Using single-particle transmission electron microscopy to reconstruct distinct populations of SgrAl oligomers, we show that in the presence of allosteric, activating DNA, the enzyme forms regular, repeating helical structures characterized by the addition of DNA-binding dimeric SgrAl subunits in a run-on manner. We also present the structure of oligomeric SgrAl at 8.6 angstrom resolution, demonstrating the conformational state of SgrAl in its activated form. Activated and oligomeric SgrAl displays key protein-protein interactions near the helix axis between its N termini, as well as allosteric protein-DNA interactions that are required for enzymatic activation. The hybrid approach reveals an unusual mechanism of enzyme activation that explains SgrAl's oligomerization and allosteric behavior.

DOI： 10.1016/j.str.2013.08.012

3DEM Loupe: analysis of macromolecular dynamics using structures from electron microscopy 査読有り

R. Nogales-Cadenas, S. Jonic, F. Tama, A. A. Arteni, D. Tabas-Madrid, M. Vazquez, A. Pascual-Montano, C. O. S. Sorzano

NUCLEIC ACIDS RESEARCH 41 巻 ( W1 ) 頁： W363 - W367 2013年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OXFORD UNIV PRESS

Electron microscopy (EM) provides access to structural information of macromolecular complexes in the 3-20 angstrom resolution range. Normal mode analysis has been extensively used with atomic resolution structures and successfully applied to EM structures. The major application of normal modes is the identification of possible conformational changes in proteins. The analysis can throw light on the mechanism following ligand binding, protein-protein interactions, channel opening and other functional macromolecular movements. In this article, we present a new web server, 3DEM Loupe, which allows normal mode analysis of any uploaded EM volume using a user-friendly interface and an intuitive workflow. Results can be fully explored in 3D through animations and movies generated by the server. The application is freely available at http://3demloupe.cnb.csic.es.

DOI： 10.1093/nar/gkt385

Consensus among multiple approaches as a reliability measure for flexible fitting into cryo-EM data 査読有り

Aqeel Ahmed, Florence Tama

JOURNAL OF STRUCTURAL BIOLOGY 182 巻 ( 2 ) 頁： 67 - 77 2013年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Cryo-electron microscopy (cryo-EM) can provide low-resolution density maps of large macromolecular assemblies. As the number of structures deposited in the Protein Data Bank by fitting a high-resolution structure into a low-resolution cryo-EM map is increasing, there is a need to revise the protocols and improve the measures for fitting. A recent study suggested using a combination of multiple automated flexible fitting approaches to improve the interpretation of cryo-EM data. The current work further explores the use of multiple approaches by validating this "consensus" fitting approach and deriving a local reliability measure. Here four different flexible fitting approaches are applied for fitting an initial structure into a simulated density map of known target structure from a dataset of proteins. It is found that the models produced from different approaches often have a consensus in conformation and are also near to the target structure, whereas cases not showing consensus are away from the target. A high correlation is also observed between the RMSF profiles calculated with respect to the average and the target structures, which indicates that the relation between consensus and accuracy can also be extended to a per-residue level. Therefore, the RMSF among the fitted models is proposed as a local reliability measure, which can be used to assess the reliability of the fit at specific regions. Hence, we encourage the community to use consensus flexible fitting with different methods to report on local reliability of the resulting models and improve the interpretation of cryo-EM data. (c) 2013 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.jsb.2013.02.002

Steered Molecular Dynamics Simulations of a Type IV Pilus Probe Initial Stages of a Force-Induced Conformational Transition 査読有り

Joseph L. Baker, Nicolas Biais, Florence Tama

PLOS COMPUTATIONAL BIOLOGY 9 巻 ( 4 ) 2013年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：PUBLIC LIBRARY SCIENCE

Type IV pili are long, protein filaments built from a repeating subunit that protrudes from the surface of a wide variety of infectious bacteria. They are implicated in a vast array of functions, ranging from bacterial motility to microcolony formation to infection. One of the most well-studied type IV filaments is the gonococcal type IV pilus (GC-T4P) from Neisseria gonorrhoeae, the causative agent of gonorrhea. Cryo-electron microscopy has been used to construct a model of this filament, offering insights into the structure of type IV pili. In addition, experiments have demonstrated that GC-T4P can withstand very large tension forces, and transition to a force-induced conformation. However, the details of force-generation, and the atomic-level characteristics of the force-induced conformation, are unknown. Here, steered molecular dynamics (SMD) simulation was used to exert a force in silico on an 18 subunit segment of GC-T4P to address questions regarding the nature of the interactions that lead to the extraordinary strength of bacterial pili. SMD simulations revealed that the buried pilin alpha 1 domains maintain hydrophobic contacts with one another within the core of the filament, leading to GC-T4P's structural stability. At the filament surface, gaps between pilin globular head domains in both the native and pulled states provide water accessible routes between the external environment and the interior of the filament, allowing water to access the pilin alpha 1 domains as reported for VC-T4P in deuterium exchange experiments. Results were also compared to the experimentally observed force-induced conformation. In particular, an exposed amino acid sequence in the experimentally stretched filament was also found to become exposed during the SMD simulations, suggesting that initial stages of the force induced transition are well captured. Furthermore, a second sequence was shown to be initially hidden in the native filament and became exposed upon stretching.

DOI： 10.1371/journal.pcbi.1003032

Molecular Model of a Soluble Guanylyl Cyclase Fragment Determined by Small-Angle X-ray Scattering and Chemical Cross-Linking 査読有り

Bradley G. Fritz, Sue A. Roberts, Aqeel Ahmed, Linda Breci, Wenzhou Li, Andrzej Weichsel, Jacqueline L. Brailey, Vicki H. Wysocki, Florence Tama, William R. Montfort

BIOCHEMISTRY 52 巻 ( 9 ) 頁： 1568 - 1582 2013年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER CHEMICAL SOC

Soluble guanylyl/guanylate cyclase (sGC) converts GTP to cGMP after binding nitric oxide, leading to smooth muscle relaxation and vasodilation. Impaired sGC activity is common in cardiovascular disease, and sGC stimulatory compounds are vigorously sought. sGC is a 150 kDa heterodimeric protein with two H-NOX domains (one with heme, one without), two PAS domains, a coiled-coil domain, and two cyclase domains. Binding of NO to the sGC heme leads to proximal histidine release and stimulation of catalytic activity. To begin to understand how binding leads to activation, we examined truncated sGC proteins from Manduca sexta (tobacco hornworm) that bind NO, CO, and stimulatory compound YC-1 but lack the cyclase domains. We determined the overall shape of truncated M. sexta sGC using analytical ultracentrifugation and small-angle X-ray scattering (SAXS), revealing an elongated molecule with dimensions of 115 angstrom x 90 angstrom x 75 angstrom. Binding of NO, CO, or YC-1 had little effect on shape. Using chemical cross-linking and tandem mass spectrometry, we identified 20 intermolecular contacts, allowing us to fit homology models of the individual domains into the SAXS-derived molecular envelope. The resulting model displays a central parallel coiled-coil platform upon which the H-NOX and PAS domains are assembled. The beta(1) H-NOX and alpha(1) PAS domains are in contact and form the core signaling complex, while the alpha(1) H-NOX domain can be removed without a significant effect on ligand binding or overall shape. Removal of 21 residues from the C-terminus yields a protein with dramatically increased proximal histidine release rates upon NO binding.

DOI： 10.1021/bi301570m

Network visualization of conformational sampling during molecular dynamics simulation 査読有り

Logan S. Ahlstrom, Joseph Lee Baker, Kent Ehrlich, Zachary T. Campbell, Sunita Patel, Ivan I. Vorontsov, Florence Tama, Osamu Miyashita

Journal of Molecular Graphics and Modelling 46 巻頁： 140 - 149 2013年

記述言語：英語掲載種別：研究論文（学術雑誌）

Effective data reduction methods are necessary for uncovering the inherent conformational relationships present in large molecular dynamics (MD) trajectories. Clustering algorithms provide a means to interpret the conformational sampling of molecules during simulation by grouping trajectory snapshots into a few subgroups, or clusters, but the relationships between the individual clusters may not be readily understood. Here we show that network analysis can be used to visualize the dominant conformational states explored during simulation as well as the connectivity between them, providing a more coherent description of conformational space than traditional clustering techniques alone. We compare the results of network visualization against 11 clustering algorithms and principal component conformer plots. Several MD simulations of proteins undergoing different conformational changes demonstrate the effectiveness of networks in reaching functional conclusions. © 2013 Elsevier Inc.

DOI： 10.1016/j.jmgm.2013.10.003

Twelve Transmembrane Helices Form the Functional Core of Mammalian MATE1 (Multidrug and Toxin Extruder 1) Protein 査読有り

Xiaohong Zhang, Xiao He, Joseph Baker, Florence Tama, Geoffrey Chang, Stephen H. Wright

JOURNAL OF BIOLOGICAL CHEMISTRY 287 巻 ( 33 ) 頁： 27971 - 27982 2012年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

The x-ray structure of the prototypic MATE family member, NorM from Vibrio cholerae, reveals a protein fold composed of 12 transmembrane helices (TMHs), confirming hydropathy analyses of the majority of (prokaryotic and plant) MATE transporters. However, the mammalian MATEs are generally predicted to have a 13th TMH and an extracellular C terminus. Here we affirm this prediction, showing that the C termini of epitope-tagged, full-length human, rabbit, and mouse MATE1 were accessible to antibodies from the extracellular face of the membrane. Truncation of these proteins at or near the predicted junction between the 13th TMH and the long cytoplasmic loop that precedes it resulted in proteins that (i) trafficked to the membrane and (ii) interacted with antibodies only after permeabilization of the plasma membrane. CHO cells expressing rbMate1 truncated at residue Gly-545 supported levels of pH-sensitive transport similar to that of cells expressing the full-length protein. Although the high transport rate of the Gly-545 truncation mutant was associated with higher levels of membrane expression (than full-length MATE1), suggesting the 13th TMH may influence substrate translocation, the selectivity profile of the mutant indicated that TMH13 has little impact on ligand binding. We conclude that the functional core of MATE1 consists of 12 (not 13) TMHs. Therefore, we used the x-ray structure of NorM to develop a homology model of the first 12 TMHs of MATE1. The model proved to be stable in molecular dynamic simulations and agreed with topology evident from preliminary cysteine scanning of intracellular versus extracellular loops.

DOI： 10.1074/jbc.M112.386979

Simulations of substrate transport in the multidrug transporter EmrD 査読有り

Joseph Baker, Stephen H. Wright, Florence Tama

PROTEINS-STRUCTURE FUNCTION AND BIOINFORMATICS 80 巻 ( 6 ) 頁： 1620 - 1632 2012年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY

EmrD is a multidrug resistance (MDR) transporter from Escherichia coli, which is involved in the efflux of amphipathic compounds from the cytoplasm, and the first MDR member of the major facilitator superfamily to be crystallized. Molecular dynamics simulation of EmrD in a phospholipid bilayer was used to characterize the conformational dynamics of the protein. Motions that support a previously proposed lateral diffusion pathway for substrate from the cytoplasmic membrane leaflet into the EmrD central cavity were observed. In addition, the translocation pathway of meta-chloro carbonylcyanide phenylhydrazone (CCCP) was probed using both standard and steered molecular dynamics simulation. In particular, interactions of a few specific residues with CCCP have been identified. Finally, a large motion of two residues, Val 45 and Leu 233, was observed with the passage of CCCP into the periplasmic space, placing a lower bound on the extent of opening required at this end of the protein for substrate transport. Overall, our simulations probe details of the transport pathway, motions of EmrD at an atomic level of detail, and offer new insights into the functioning of MDR transporters. Proteins 2012; (c) 2012 Wiley Periodicals, Inc.

DOI： 10.1002/prot.24056

Twelve transmembrane helices form the functional core of mammalian MATE1 査読有り

Zhang Xiaohong, He Xiao, Baker Joseph, Tama Florence, Chang Geoffrey, Wright Stephen

FASEB JOURNAL 26 巻頁： . 2012年4月

Consensus among flexible fitting approaches improves the interpretation of cryo-EM data 査読有り

Aqeel Ahmed, Paul C. Whitford, Karissa Y. Sanbonmatsu, Florence Tama

JOURNAL OF STRUCTURAL BIOLOGY 177 巻 ( 2 ) 頁： 561 - 570 2012年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Cryo-elecron microscopy (cryo-EM) can provide important structural information of large macromolecular assemblies in different conformational states. Recent years have seen an increase in structures deposited in the Protein Data Bank (PDB) by fitting a high-resolution structure into its low-resolution cryo-EM map. A commonly used protocol for accommodating the conformational changes between the X-ray structure and the cryo-EM map is rigid body fitting of individual domains. With the emergence of different flexible fitting approaches, there is a need to compare and revise these different protocols for the fitting. We have applied three diverse automated flexible fitting approaches on a protein dataset for which rigid domain fitting (RDF) models have been deposited in the PDB. In general, a consensus is observed in the conformations, which indicates a convergence from these theoretically different approaches to the most probable solution corresponding to the cryo-EM map. However, the result shows that the convergence might not be observed for proteins with complex conformational changes or with missing densities in cryo-EM map. In contrast, RDF structures deposited in the PDB can represent conformations that not only differ from the consensus obtained by flexible fitting but also from X-ray crystallography. Thus, this study emphasizes that a "consensus" achieved by the use of several automated flexible fitting approaches can provide a higher level of confidence in the modeled configurations. Following this protocol not only increases the confidence level of fitting, but also highlights protein regions with uncertain fitting. Hence, this protocol can lead to better interpretation of cryo-EM data. (C) 2011 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.jsb.2011.10.002

Phosphorylated Smooth Muscle Heavy Meromyosin Shows an Open Conformation: Implications for the Structure of Myosin with One Head Phosphorylated 査読有り

Taylor Kenneth A., Baumann Bruce A. J., Taylor Dianne W., Huang Zhong, Tama Florence, Fagnant Patricia M., Trybus Kathleen M.

BIOPHYSICAL JOURNAL 102 巻 ( 3 ) 頁： 159A-159A 2012年1月

Phosphorylated Smooth Muscle Heavy Meromyosin Shows an Open Conformation Linked to Activation 査読有り

Bruce A. J. Baumann, Dianne W. Taylor, Zhong Huang, Florence Tama, Patricia M. Fagnant, Kathleen M. Trybus, Kenneth A. Taylor

JOURNAL OF MOLECULAR BIOLOGY 415 巻 ( 2 ) 頁： 274 - 287 2012年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD

Smooth muscle myosin and smooth muscle heavy meromyosin (smHMM) are activated by regulatory light chain phosphorylation, but the mechanism remains unclear. Dephosphorylated, inactive smHMM assumes a closed conformation with asymmetric intramolecular head head interactions between motor domains. The "free head" can bind to actin, but the actin binding interface of the "blocked head" is involved in interactions with the free head. We report here a three-dimensional structure for phosphorylated, active smHMM obtained using electron crystallography of two-dimensional arrays. Head head interactions of phosphorylated smHMM resemble those found in the dephosphorylated state but occur between different molecules, not within the same molecule. The light chain binding domain structure of phosphorylated smHMM differs markedly from that of the "blocked" head of dephosphorylated smHMM. We hypothesize that regulatory light chain phosphorylation opens the inhibited conformation primarily by its effect on the blocked head. Singly phosphorylated smHMM is not compatible with the closed conformation if the blocked head is phosphorylated. This concept has implications for the extent of myosin activation at low levels of phosphorylation in smooth muscle. (C) 2011 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.jmb.2011.10.047

Excited states of ribosome translocation revealed through integrative molecular modeling 査読有り

Paul C. Whitford, Aqeel Ahmed, Yanan Yu, Scott P. Hennelly, Florence Tama, Christian M. T. Spahn, Jose N. Onuchic, Karissa Y. Sanbonmatsu

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 108 巻 ( 47 ) 頁： 18943 - 18948 2011年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATL ACAD SCIENCES

The dynamic nature of biomolecules leads to significant challenges when characterizing the structural properties associated with function. While X-ray crystallography and imaging techniques (such as cryo-electron microscopy) can reveal the structural details of stable molecular complexes, strategies must be developed to characterize configurations that exhibit only marginal stability (such as intermediates) or configurations that do not correspond to minima on the energy landscape (such as transition-state ensembles). Here, we present a methodology (MDfit) that utilizes molecular dynamics simulations to generate configurations of excited states that are consistent with available biophysical and biochemical measurements. To demonstrate the approach, we present a sequence of configurations that are suggested to be associated with transfer RNA (tRNA) movement through the ribosome (translocation). The models were constructed by combining information from X-ray crystallography, cryo-electron microscopy, and biochemical data. These models provide a structural framework for translocation that may be further investigated experimentally and theoretically to determine the precise energetic character of each configuration and the transition dynamics between them.

DOI： 10.1073/pnas.1108363108

Structure modeling from small angle X-ray scattering data with elastic network normal mode analysis 査読有り

Osamu Miyashita, Christian Gorba, Florence Tama

JOURNAL OF STRUCTURAL BIOLOGY 173 巻 ( 3 ) 頁： 451 - 460 2011年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Computational algorithms to construct structural models from SAXS experimental data are reviewed. SAXS data provides a wealth of information to study the structure and dynamics of biological molecules, however it does not provide atomic details of structures. Thus combining the low-resolution data with already known X-ray structure is a common approach to study conformational transitions of biological molecules. This review provides a survey of SAXS modeling approaches. In addition, we will discuss theoretical backgrounds and performance of our approach, in which elastic network normal mode analysis is used to predict reasonable conformational transitions from known X-ray structures, and find alternative conformations that are consistent with SAXS data. (C) 2010 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.jsb.2010.09.008

Normal Mode Flexible Fitting of High-Resolution Structures of Biological Molecules Toward SAXS Data. 国際誌

Christian Gorba, Florence Tama

Bioinformatics and biology insights 4 巻頁： 43 - 54 2010年6月

記述言語：英語掲載種別：研究論文（学術雑誌）

We present a method to reconstruct a three-dimensional protein structure from an atomic pair distribution function derived from the scattering intensity profile from SAXS data by flexibly fitting known x-ray structures. This method uses a linear combination of low-frequency normal modes from an elastic network description of the molecule in an iterative manner to deform the structure to conform optimally to the target pair distribution function derived from SAXS data. For computational efficiency, the protein and water molecules included in the protein first hydration shell are coarse-grained. In this paper, we demonstrate the validity of our coarse-graining approach to study SAXS data. Illustrative results of our flexible fitting studies on simulated SAXS data from five different proteins are presented.

Three-dimensional structure of the anthrax toxin pore inserted into lipid nanodiscs and lipid vesicles 査読有り

H. Katayama, J. Wang, F. Tama, L. Chollet, E. P. Gogol, R. J. Collier, M. T. Fisher

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 107 巻 ( 8 ) 頁： 3453 - 3457 2010年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATL ACAD SCIENCES

A major goal in understanding the pathogenesis of the anthrax bacillus is to determine how the protective antigen (PA) pore mediates translocation of the enzymatic components of anthrax toxin across membranes. To obtain structural insights into this mechanism, we constructed PA-pore membrane complexes and visualized them by using negative-stain electron microscopy. Two populations of PA pores were visualized in membranes, vesicle-inserted and nanodisc-inserted, allowing us to reconstruct two virtually identical PA-pore structures at 22-angstrom resolution. Reconstruction of a domain 4-truncated PA pore inserted into nanodiscs showed that this domain does not significantly influence pore structure. Normal mode flexible fitting of the x-ray crystallographic coordinates of the PA prepore indicated that a prominent flange observed within the pore lumen is formed by the convergence of mobile loops carrying Phe427, a residue known to catalyze protein translocation. Our results have identified the location of a crucial functional element of the PA pore and documented the value of combining nanodisc technology with electron microscopy to examine the structures of membrane-interactive proteins.

DOI： 10.1073/pnas.1000100107

Biased coarse-grained molecular dynamics simulation approach for flexible fitting of X-ray structure into cryo electron microscopy maps 査読有り

Ivan Grubisic, Maxim N. Shokhirev, Marek Orzechowski, Osamu Miyashita, Florence Tama

JOURNAL OF STRUCTURAL BIOLOGY 169 巻 ( 1 ) 頁： 95 - 105 2010年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Several approaches have been introduced to interpret, in terms of high-resolution structure, low-resolution structural data as obtained from cryo-EM. As conformational changes are often observed in biological molecules, these techniques need to take into account the flexibility of proteins. Flexibility has been described in terms of movement between rigid domains and between rigid secondary structure elements, which present some limitations for studying dynamical properties. Normal mode analysis has also been used, but is limited to medium resolution data. All-atom molecular dynamics fitting techniques are more appropriate to fit structures into higher-resol uti on data as full protein flexibility is considered, but are cumbersome in terms of computational time. Here, we introduce a coarse-grained approach; a Go-model was used to represent biological molecules, combined with biased molecular dynamics to reproduce accurately conformational transitions. Illustrative examples on simulated data are shown. Accurate fittings can be obtained for resolution ranging from 5 to 20 angstrom. The approach was also tested on experimental data of Elongation Factor G and Escherichia coli RNA polymerase, where its validity is compared to previous models obtained from different techniques. This comparison demonstrates that quantitative flexible techniques, as opposed to manual docking, need to be considered to interpret low-resolution data. (C) 2009 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.jsb.2009.09.010

Flexible Fitting of High-Resolution X-Ray Structures into Cryoelectron Microscopy Maps Using Biased Molecular Dynamics Simulations 査読有り

Marek Orzechowski, Florence Tama

BIOPHYSICAL JOURNAL 95 巻 ( 12 ) 頁： 5692 - 5705 2008年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：CELL PRESS

A methodology for flexible fitting of all-atom high-resolution structures into low-resolution cryoelectron microscopy (cryo-EM) maps is presented. Flexibility of the modeled structure is simulated by classical molecular dynamics and an additional effective potential is introduced to enhance the fitting process. The additional potential is proportional to the correlation coefficient between the experimental cryo-EM map and a synthetic map generated for an all-atom structure being fitted to the map. The additional forces are calculated as a gradient of the correlation coefficient. During the molecular dynamics simulations under the additional forces, the molecule undergoes a conformational transition that maximizes the correlation coefficient, which results in a high-accuracy fit of all-atom structure into a cryo-EM map. Using five test proteins that exhibit structural rearrangement during their biological activity, we demonstrate performance of our method. We also test our method on the experimental cryo-EM of elongation factor G and show that the model obtained is comparable to previous studies. In addition, we show that over fitting can be avoided by assessing the quality of the fitted model in terms of correlation coefficient and secondary structure preservation.

DOI： 10.1529/biophysj.108.139451

Normal-mode flexible fitting of high-resolution structure of biological molecules toward one-dimensional low-resolution data 査読有り

Christian Gorba, Osamu Miyashita, Florence Tama

BIOPHYSICAL JOURNAL 94 巻 ( 5 ) 頁： 1589 - 1599 2008年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：BIOPHYSICAL SOC

We present a method for reconstructing a 3D structure from a pair distribution function by flexibly fitting known x-ray structures toward a conformation that agrees with the low-resolution data. This method uses a linear combination of low-frequency normal modes from elastic-network description of the molecule in an iterative manner to deform the structure optimally to conform to the target pair distribution function. A simple function, pair distance distribution function between atoms, is chosen as a test model to establish computational algorithms-optimization algorithm and scoring function-that can utilize low-resolution 1 D data. To select a correct structural model based on less information, we developed a scoring function that takes into account a characteristic of pair distribution functions. In addition, we employ a new optimization algorithm, the trusted region method, that relies on both first and second derivatives of the scoring function. Illustrative results of our studies on simulated 1D data from five different proteins, for which large conformational changes are known to occur, are presented.

DOI： 10.1529/biophysj.107.122218

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