Faculty Profiles - TAMA Florence

Papers - TAMA Florence

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Editorial overview: Macromolecular assemblies Reviewed

Toshiyuki Shimizu, Florence Tama

CURRENT OPINION IN STRUCTURAL BIOLOGY Vol. 43 page： VII - ix 2017.4

Hybrid Approach for Biological Structure Modeling: Cryo-electron Microscopy and X-ray Free Electron Laser Data Open Access

MIYASHITA Osamu, TAMA Florence

Seibutsu Butsuri Vol. 57 ( 2 ) page： 90 - 94 2017

Language：Japanese Publisher：The Biophysical Society of Japan General Incorporated Association

<p>Hybrid approaches combine computational modeling techniques with low-resolution structural data. Such approaches have proven to be powerful tools to obtain new 3D structural and dynamical information on biological systems. Currently, major applications focus on cryo-EM data. Methods and some applications to construct atomic structural models of new functional states will be reviewed. In addition, possible extension to data from X-ray free electron laser, which currently provides low-resolution data, will be discussed.</p>

DOI： 10.2142/biophys.57.090

Local thermodynamics of the water molecules around single- and double-stranded DNA studied by grid inhomogeneous solvation theory Reviewed

Miki Nakano, Hisae Tateishi-Karimata, Shigenori Tanaka, Florence Tama, Osamu Miyashita, Shu-ichi Nakano, Naoki Sugimoto

CHEMICAL PHYSICS LETTERS Vol. 660 page： 250 - 255 2016.9

Language：Japanese Publishing type：Research paper (scientific journal) Publisher：ELSEVIER SCIENCE BV

Thermodynamic properties of water molecules around single- and double-stranded DNAs (ssDNAs and dsDNAs) with different sequences were investigated using grid inhomogeneous solvation theory. Free energies of water molecules solvating the minor groove of dsDNAs are lower than those near ssDNAs, while water molecules should be released during the formation of dsDNA. Free energies of water molecules around dsDNA are lower than those around ssDNA even in the second and third hydration shells. Our findings will help to clarify the role of water molecules in the formation of dsDNA from ssDNAs, thus facilitating the designs of drugs or nanomaterials using DNA. (C) 2016 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.cplett.2016.08.032

Hybrid approach for structural modeling of biological systems from X-ray free electron laser diffraction patterns Reviewed

Atsushi Tokuhisa, Slavica Jonic, Florence Tama, Osamu Miyashita

JOURNAL OF STRUCTURAL BIOLOGY Vol. 194 ( 3 ) page： 325 - 336 2016.6

Language：Japanese Publishing type：Research paper (scientific journal) Publisher：ACADEMIC PRESS INC ELSEVIER SCIENCE

We present a new hybrid approach for structural modeling using X-ray free electron laser (XFEL) diffraction patterns from non-crystalline biological samples. Reconstruction of a 3D structure requires a large number of diffraction patterns; however, in the current XFEL experiments with biological systems, the analysis often relies on a small number of 2D diffraction patterns. In this study, we explore the strategies to identify plausible 3D structural models by combining the 2D analysis of such diffraction patterns with computational modeling (normal mode analysis or molecular dynamics simulations). As the first step toward such hybrid modeling, we established a protocol to assess the agreement between the model structure and the target XFEL diffraction pattern and showed that XFEL data can be used to study the conformational transitions of biological molecules. We tested the proposed algorithms using data of three biomolecular complexes of different sizes (elongation factor 2, CCM virus, and ribosome) and examined the experimental conditions that are required to perform such studies, in particular the XFEL beam intensity requirements. The results indicate that the current beam intensity is close to a strength that enables us to study conformational transitions of macromolecules, such as ribosomes. The proposed algorithm can be combined with molecular mechanics approaches, such as molecular dynamics simulations and normal mode analysis, to generate a large number of candidate structures to perform hybrid structural modeling. (C) 2016 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.jsb.2016.03.009

Thermodynamic properties of water molecules in the presence of cosolute depend on DNA structure: a study using grid inhomogeneous solvation theory Reviewed Open Access

Nakano Miki, Tateishi-Karimata Hisae, Tanaka Shigenori, Tama Florence, Miyashita Osamu, Nakano Shu-ichi, Sugimoto Naoki

NUCLEIC ACIDS RESEARCH Vol. 43 ( 21 ) page： 10114-10125 2015.12

Hybrid Electron Microscopy Normal Mode Analysis graphical interface and protocol Reviewed

Carlos Oscar S. Sorzano, Jose Miguel de la Rosa-Trevin, Florence Tama, Slavica Jonic

JOURNAL OF STRUCTURAL BIOLOGY Vol. 188 ( 2 ) page： 134 - 141 2014.11

Language：English Publishing type：Research paper (scientific journal) Publisher：ACADEMIC PRESS INC ELSEVIER SCIENCE

This article presents an integral graphical interface to the Hybrid Electron Microscopy Normal Mode Analysis (HEMNMA) approach that was developed for capturing continuous motions of large macromolecular complexes from single-particle EM images. HEMNMA was shown to be a good approach to analyze multiple conformations of a macromolecular complex but it could not be widely used in the EM field due to a lack of an integral interface. In particular, its use required switching among different software sources as well as selecting modes for image analysis was difficult without the graphical interface. The graphical interface was thus developed to simplify the practical use of HEMNMA. It is implemented in the open-source software package Xmipp 3.1 (http://xmipp.cnb.csic.es) and only a small part of it relies on MATLAB that is accessible through the main interface. Such integration provides the user with an easy way to perform the analysis of macromolecular dynamics and forms a direct connection to the single-particle reconstruction process. A step-by-step HEMNMA protocol with the graphical interface is given in full details in Supplementary material. The graphical interface will be useful to experimentalists who are interested in studies of continuous conformational changes of macromolecular complexes beyond the modeling of continuous heterogeneity in single particle reconstruction. (C) 2014 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.jsb.2014.09.005

Replica Exchange Molecular Dynamics Simulations Provide Insight into Substrate Recognition by Small Heat Shock Proteins Reviewed Open Access

Sunita Patel, Elizabeth Vierling, Florence Tama

BIOPHYSICAL JOURNAL Vol. 106 ( 12 ) page： 2644 - 2655 2014.6

Language：English Publishing type：Research paper (scientific journal) Publisher：CELL PRESS

The small heat shock proteins (sHSPs) are a virtually ubiquitous and diverse group of molecular chaperones that can bind and protect unfolding proteins from irreversible aggregation. It has been suggested that intrinsic disorder of the N-terminal arm (NTA) of sHSPs is important for substrate recognition. To investigate conformations of the NTA that could recognize substrates we performed replica exchange molecular dynamics simulations. Behavior at normal and stress temperatures of the dimeric building blocks of dodecameric HSPs from wheat (Ta16.9) and pea (Ps18.1) were compared because they display high sequence similarity, but Ps18.1 is more efficient in binding specific substrates. In our simulations, the NTAs of the dimer are flexible and dynamic; however, rather than exhibiting highly extended conformations they retain considerable alpha-helical character and contacts with the conserved alpha-crystallin domain (ACD). Network analysis and clustering methods reveal that there are two major conformational forms designated either "open" or "closed" based on the relative position of the two NTAs and their hydrophobic solvent accessible surface area. The equilibrium constant for the closed to open transition is significantly different for Ta16.9 and Ps18.1, with the latter showing more open conformations at elevated temperature correlated with its more effective chaperone activity. In addition, the Ps18.1 NTAs have more hydrophobic solvent accessible surface than those of Ta16.9. NTA hydrophobic patches are comparable in size to the area buried in many protein-protein interactions, which would enable sHSPs to bind early unfolding intermediates. Reduced interactions of the Ps18.1 NTAs with each other and with the ACD contribute to the differences in dynamics and hydrophobic surface area of the two sHSPs. These data support a major role for the conformational equilibrium of the NTA in substrate binding and indicate features of the NTA that contribute to sHSP chaperone efficiency.

DOI： 10.1016/j.bpj.2014.04.048

Macromolecular structures probed by combining single-shot free-electron laser diffraction with synchrotron coherent X-ray imaging Reviewed Open Access

Marcus Gallagher-Jones, Yoshitaka Bessho, Sunam Kim, Jaehyun Park, Sangsoo Kim, Daewoong Nam, Chan Kim, Yoonhee Kim, Do Young Noh, Osamu Miyashita, Florence Tama, Yasumasa Joti, Takashi Kameshima, Takaki Hatsui, Kensuke Tono, Yoshiki Kohmura, Makina Yabashi, S. Samar Hasnain, Tetsuya Ishikawa, Changyong Song

NATURE COMMUNICATIONS Vol. 5 2014.5

Language：English Publishing type：Research paper (scientific journal) Publisher：NATURE PUBLISHING GROUP

Nanostructures formed from biological macromolecular complexes utilizing the self-assembly properties of smaller building blocks such as DNA and RNA hold promise for many applications, including sensing and drug delivery. New tools are required for their structural characterization. Intense, femtosecond X-ray pulses from X-ray free-electron lasers enable single-shot imaging allowing for instantaneous views of nanostructures at ambient temperatures. When combined judiciously with synchrotron X-rays of a complimentary nature, suitable for observing steady-state features, it is possible to perform ab initio structural investigation. Here we demonstrate a successful combination of femtosecond X-ray single-shot diffraction with an X-ray free-electron laser and coherent diffraction imaging with synchrotron X-rays to provide an insight into the nanostructure formation of a biological macromolecular complex: RNA interference microsponges. This newly introduced multimodal analysis with coherent X-rays can be applied to unveil nano-scale structural motifs from functional nanomaterials or biological nanocomplexes, without requiring a priori knowledge.

DOI： 10.1038/ncomms4798

Iterative Elastic 3D-to-2D Alignment Method Using Normal Modes for Studying Structural Dynamics of Large Macromolecular Complexes Reviewed Open Access

Qiyu Jin, Carlos Oscar S. Sorzano, Jose Miguel de la Rosa-Trevin, Jose Roman Bilbao-Castro, Rafael Nunez-Ramirez, Oscar Llorca, Florence Tama, Slavica Jonic

STRUCTURE Vol. 22 ( 3 ) page： 496 - 506 2014.3

Language：English Publishing type：Research paper (scientific journal) Publisher：CELL PRESS

This article presents a method to study large-scale conformational changes by combining electron microscopy (EM) single-particle image analysis and normal mode analysis (NMA). It is referred to as HEMNMA, which stands for hybrid electron microscopy normal mode analysis. NMA of a reference structure (atomic-resolution structure or EM volume) is used to predict possible motions that are then confronted with EM images within an automatic iterative elastic 3D-to-2D alignment procedure to identify actual motions in the imaged samples. HEMNMA can be used to extensively analyze the conformational changes and may be used in combination with classic discrete procedures. The identified conformations allow modeling of deformation pathways compatible with the experimental data. HEMNMA was tested with synthetic and experimental data sets of E. coli 70S ribosome, DNA polymerase Pol alpha and B subunit complex of the eukaryotic primosome, and tomato bushy stunt virus.

DOI： 10.1016/j.str.2014.01.004

ELASTIC IMAGE REGISTRATION TO FULLY EXPLORE MACROMOLECULAR DYNAMICS BY ELECTRON MICROSCOPY Reviewed

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

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

ELASTIC IMAGE REGISTRATION TO FULLY EXPLORE MACROMOLECULAR DYNAMICS BY ELECTRON MICROSCOPY Reviewed

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

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

Language：English Publishing type：Research paper (international conference proceedings) Publisher：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 Reviewed Open Access

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

STRUCTURE Vol. 21 ( 10 ) page： 1848 - 1858 2013.10

Language：English Publishing type：Research paper (scientific journal) Publisher：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 Reviewed Open Access

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 Vol. 41 ( W1 ) page： W363 - W367 2013.7

Language：English Publishing type：Research paper (scientific journal) Publisher：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 Reviewed

Aqeel Ahmed, Florence Tama

JOURNAL OF STRUCTURAL BIOLOGY Vol. 182 ( 2 ) page： 67 - 77 2013.5

Language：English Publishing type：Research paper (scientific journal) Publisher：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 Reviewed Open Access

Joseph L. Baker, Nicolas Biais, Florence Tama

PLOS COMPUTATIONAL BIOLOGY Vol. 9 ( 4 ) 2013.4

Language：English Publishing type：Research paper (scientific journal) Publisher：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 Reviewed Open Access

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 Vol. 52 ( 9 ) page： 1568 - 1582 2013.3

Language：English Publishing type：Research paper (scientific journal) Publisher：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 Reviewed Open Access

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 Vol. 46 page： 140 - 149 2013

Language：English Publishing type：Research paper (scientific journal)

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 Reviewed Open Access

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

JOURNAL OF BIOLOGICAL CHEMISTRY Vol. 287 ( 33 ) page： 27971 - 27982 2012.8

Language：English Publishing type：Research paper (scientific journal) Publisher：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 Reviewed Open Access

Joseph Baker, Stephen H. Wright, Florence Tama

PROTEINS-STRUCTURE FUNCTION AND BIOINFORMATICS Vol. 80 ( 6 ) page： 1620 - 1632 2012.6

Language：English Publishing type：Research paper (scientific journal) Publisher：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 Reviewed

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

FASEB JOURNAL Vol. 26 page： . 2012.4

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