Scientific Program

Conference Series Ltd invites all the participants across the globe to attend 9th International Conference on Structural Biology Zurich, Switzerland.

Day 1 :

Structural Biology 2017 International Conference Keynote Speaker Henry M Sobell photo
Biography:

Henry M. Sobell completed his studies at Brooklyn Technical High School (1948-1952), Columbia College (1952-1956), and the University of Virginia School of Medicine (1956-1960).  Instead of practicing clinical medicine, he then went to the Massachusetts Institute of Technology (MIT) to join Professor Alexander Rich in the Department of Biology (1960-1965), where, as a Helen Hay Whitney Postdoctoral Fellow, he learned the technique of single crystal X-ray analysis.  He then joined the Chemistry Department at the University of Rochester, having been subsequently jointly appointed to both the Chemistry and Molecular Biophysics departments (the latter at the University of Rochester School of Medicine and Dentistry), becoming a full tenured Professor in both departments (1965-1993).  He is now retired and living in the Adirondacks in New York, USA.

Abstract:

Premeltons are examples of emergent structures (i.e., structural solitons) that arise spontaneously in DNA due to the presence of nonlinear excitations in its structure.  They are of two kinds: B-B (or A-A) premeltons form at specific DNA-regions to nucleate site-specific DNA melting.  These are stationary and, being globally nontopological, undergo breather motions that allow drugs and dyes to intercalate into DNA.  B-A (or A-B) premeltons, on the other hand, are mobile, and being globally topological, act as phase-boundaries transforming B- into A- DNA during the structural phase-transition.  They are not expected to undergo breather-motions.  A key feature of both types of premeltons is the presence of an intermediate structural-form in their central regions (proposed as being a transition-state intermediate in DNA-melting and in the B- to A- transition), which differs from either A- or B- DNA. Called beta-DNA, this is both metastable and hyperflexible – and contains an alternating sugar-puckering pattern along the polymer-backbone combined with the partial-unstacking (in its lower energy-forms) of every other base-pair.  Beta-DNA is connected to either B- or to A- DNA on either side by boundaries possessing a gradation of nonlinear structural-change, these being called the kink and the antikink regions.  The presence of premeltons in DNA leads to a unifying theory to understand much of DNA physical-chemistry and molecular-biology.  In particular, premeltons are predicted to define the 5’ and 3’ ends of genes in naked-DNA and DNA in active-chromatin, this having important implications for understanding physical aspects of the initiation, elongation and termination of RNA-synthesis during transcription.  For these and other reasons, the model will be of broader interest to the general audience working in these areas.  The model explains a wide variety of data, and carries within it a number of experimental predictions – all readily testable – as will be described in my talk.

Keynote Forum

Tilman Schirmer

University of Basel, Switzerland

Keynote: Structural biology of c-di-GMP mediated signaling
Structural Biology 2017 International Conference Keynote Speaker Tilman Schirmer photo
Biography:

Abstract:

In addition to the well-known cyclic nucleotides, cAMP and cGMP, bacteria utilize cyclic di-guanosine monophosphate (c-di-GMP) to control various cellular processes. Hereby, the cellular level of the messenger is set by the antagonistic activities of diguanylate cyclases and specific phosphodiesterases. In a given organism, there are usually multiple variants of the two enzymes, which are tightly regulated by a variety of external and internal cues due to the presence of specialized sensory or regulatory domains. Fundamental cellular processes, such as bacterial life style, biofilm formation, and cell cycle control are thus getting controlled in a coordinated fashion by downstream c-di-GMP receptors in response to the input signals.

Crystal structures in combination with biochemical and biophysical analyses reveal that both GGDEF diguanylate cyclase and EAL phosphodiesterase domains are active only as homotypic dimers. In the full-length enzymes, attainment of the competent quarternary structure depends on the signaling state of the accessory domains (e.g. Rec, PAS, GAF), that typically dimerize or change their dimeric structure upon signal perception. Histidine kinases and transcription factors use very similar regulatory domains to control output function in a dimeric context. It can be inferred that the modular arrangement of catalytic and regulatory dimers, both forming homotypic interactions, facilitates their recombination during evolution.

As an example for c-di-GMP mediated allosteric control of a downstream effector, the effect of c-di-GMP binding to the bifunctional histidine kinase CckA from C. crescentus will be presented. It was found that c-di-GMP promotes the phosphatase activity of the enzyme via stabilization of the phosphatase competent constellation due to non-covalent domain cross-linking. In silico analyses predict that c-di-GMP control is widespread among bacterial histidine kinases, arguing that it can replace or modulate canonical transmembrane signaling.

  • Track 1: 3D Structure Determination | Track 2: Computational Approaches in Structural Biology | Track 3: Structural Biology and Single Molecules
Location:

Session Introduction

Beat Vögeli

University of Colorado at Denver, USA

Title: Functional protein conformation networks probed by NMR nanorulers
Biography:

Beat Vögeli has his expertise in nuclear magnetic resonance (NMR) spectroscopy of biomacromolecules. He develops methodology for the elucidation of conformation and communication networks within and between proteins and nucleic acids. He received his Ph.D. degree at the ETH Zürich in the group of Konstantin Pervushin. After a postdoctoral stay at the National Institutes of Health, Bethesda USA, in the group of Ad Bax, he returned to ETH Zürich to become Oberassistant in the group of Roland Riek and Privatdozent. He is currently an assistant professor at the University of Colorado at Denver in the Department of Biochemistry and Molecular Genetics.

Abstract:

Beat Vögeli has his expertise in nuclear magnetic resonance (NMR) spectroscopy of biomacromolecules. He develops methodology for the elucidation of conformation and communication networks within and between proteins and nucleic acids. He received his Ph.D. degree at the ETH Zürich in the group of Konstantin Pervushin. After a postdoctoral stay at the National Institutes of Health, Bethesda USA, in the group of Ad Bax, he returned to ETH Zürich to become Oberassistant in the group of Roland Riek and Privatdozent. He is currently an assistant professor at the University of Colorado at Denver in the Department of Biochemistry and Molecular Genetics.

Biography:

Chromatin is the ubiquitous protein-DNA complex that forms the structural basis of DNA condensation in all eukaryotic organisms. Packaging and depackaging of chromatin, called chromatin remodeling, plays a central role in all cellular processes that involve chromosomes such as transcription, replication, recombination and repair. Detailed knowledge of the principles and mechanisms underlying this control of DNA condensation is thus vital for understanding many diseases, including neurological disorders and cancer. The physical mechanisms governing these processes however, are still largely unknown. I am interested in developing and using modern biophysical techniques to unravel the physics behind DNA condensation and its role in transcription regulation.

Abstract:

The folding of chromatin defines access to our genes and therefore plays a pivotal role in transcription regulation. However, the structure of chromatin fibers is poorly defined and heavily debated. We used single-molecule techniques to probe and manipulate the dynamics of nucleosomes in individual chromatin fibers. These novel methods were initially applied to synthetic, highly homogeneous nucleosomal arrays and yielded unprecedented insight in the structure and dynamics of chromatin.
 
With single pair Forster Resonance Energy Transfer we showed that the nucleosome is very dynamic, unwrapping half of its DNA four times per second. Using single molecule force spectroscopy, it was possible to measure the kinetics of this unfolding, both in single nucleosomes and in well-defined arrays of nucleosomes that fold into a 30 nm fiber. Analysis of the unfolding pattern reveals a linker length dependence of the higher order folding.
The linker length in vivo however varies, and to obtain insight the positioning of nucleosomes we developed a simple statistical physics model that captures sequence dependent positioning effects for both reconstitutions on synthetic DNA and chromatin in vivo.
 
We recently developed a method to purify specific chromatin fragments from yeast without crosslinking the fiber while maintaining the complexity that provides functionality to our epi-genome. I will show the first single-molecule force spectroscopy results on intact, native fibers which uniquely probe chromatin structure, composition and variations in it at the single-molecule level.

Biography:

Milton Saier has broad interests in molecular biology. He directs the research in two laboratories, one a bioinformatics laboratory, and the other a molecular genetics/biochemistry lab. His interests deal with mechanisms and evolution of transport protein functions and genomic analyses of transport systems in microorganisms (dry lab). In the wet lab, mechanisms of transcriptional and enzyme activity regulation are the focus of their research. His current efforts deal with insertion sequence-mediated directed mutation control by DNA configuration and DNA binding proteins in E. coli. The present abstract results from efforts of himself and his research associate, Dr. Zhongge Zhang, as well as others. Recent efforts in the biochemistry section of the lab have revealed a novel mechanism of global regulation of carbon and energy metabolism. Saier has published over 700 articles over the past 40 years on a wide range of subjects including molecular biological and environmental topics.

Abstract:

Our laboratory maintains and curates the Transporter Classification Database (TCDB, www.tcdb.org), which is continually updated and expanded, and currently includes over 11,000 transport systems, 1000 families and 64 superfamilies, citing over 12,500 references. We have developed software that allow us to identify distant relationships between proteins using the Transitivity Rule with results confirmed by sequence motif similarities, Pfam assignments, common evolutionary pathways and 3-D structural comparisons. Many superfamilies contain proteins of only one mechanistic type (i.e., channel proteins, secondary carriers, primary active transporters or group transporters (TC classes 1-4, respectively)). However, our recent analyses show that some superfamilies include multiple protein functional types. For example, the Transporter/Opsin/G-protein (TOG) superfamily includes channels, carriers, primary active transporters and receptors, but, no enzymes. Another superfamily, the 4 TMS junctional complex (4JC) superfamily includes gap and tight junctional complex proteins, simple channels and channel auxiliary proteins, but no carriers or primary active transporters. The Major Facilitator Superfamily (MFS) consists largely of secondary carriers, lacks channel proteins, but includes transmembrane domains in P-type ATPases, integral membrane proteases, glycosyltransferases and tRNA synthases. In some cases we could show that within a single superfamily, 3-D structures can vary so much that their common features become unrecognizable, even though primary sequence similarity is appreciable. These experimental approaches will be discussed and compared.

CongBao Kang

Agency for Science, Technology and Research (A*STAR), Singapore

Title: Structural and dynamic studies of DENV and ZIKV proteases and its insight into inhibitor design
Biography:

CongBao Kang received his Ph.D. from School of Biological Sciences at Nanyang Technological University (NTU). He was a research fellow at Centre for Structural Biology, Vanderbilt University, where he was working on structural determination of disease-related membrane proteins. He is currently the group leader of high End NMR group at ETC. His group is working on protein structure, dynamics and its interaction with potential drug candidates using solution NMR spectroscopy. The goal of his group is to provide structural information of a target protein to the medicine chemists to understand structure-activity relationship of potent compounds. His group is involving in hits identification, hits to lead, and lead optimization steps of the drug discovery process. His is currently working on target-based drug discoveries. The targets include methyltransferases, kinases, ion channels, membrane-bound receptors, protein-protein interactions, and viral proteins.

Abstract:

Dengue virus (DENV and Zika virus (ZIKV) belong to Flaviviridae genus which contains important human pathogens. DENV affects people living in tropical and subtropical regions. DENV infection can cause serious diseases such as dengue fever. ZIKV has drawn worldwide attention because of the outbreak in 2015. Viral genome of a flavivirus encodes a polyprotein that can be processed into structural and non-structural (NS) proteins by both host and viral proteases. Viral protease is a two-component serine protease formed by a cofactor region (~40 aa) from NS2B and a protease region (~170 aa) from NS3. The NS2B-NS3 protease of DENV or ZIKV is a validated target because of their function in maturation of viral proteins. Structural studies have been conducted for both DENV and ZIKV proteases. For DENV, previous studies have demonstrated that the free protease adopts an open conformation in which the C-terminal part of the NS2B cofactor region stays away from the active site. In the presence of an inhibitor, DENV protease forms a closed conformation in which the C-terminal region of NS2B forms part of the active site and interacts with the inhibitor. Our NMR study reveals that an unlinked DENV protease adopts the closed conformation in solution. Based on the knowledge on DENV protease, several constructs were made for ZIKV protease. Structural studies demonstrated that ZIKV protease adopts the closed conformation in the absence and presence of an inhibitor or substrate. The linker or enzymatic cleavage site present between NS2B and NS3 may affect inhibitor to interact with the active site. Our accumulated studies have shown that the unlinked protease construct can be used for studying protease-inhibitor interactions. We have demonstrated that the unlinked ZIKV protease interacts with different types of inhibitors. Our studies will be helpful for structure-based inhibitor design against both ZIKV and DENV proteases. 

Biography:

The Kolaczkowski Lab uses a combination of computational biology, statistics, structural modeling and biochemistry to examine how molecular systems evolve. We are particularly interested in the evolution of innate immunity in animals and plants. Dr. Kolaczkowski’s research program focuses on developing cutting-edge tools for the characterization of molecular-evolutionary processes and applying these approaches to learn how long-term protein evolution works. Dr. Kolaczkowski’s recent work has advanced the fields of ancestral sequence resurrection (ASR) and protein function prediction while shedding new light on the evolution of innate antiviral immunity and RNA interference.

Abstract:

Determining the evolutionary forces driving transitions in protein function, the structural mechanisms through which functions change and the effects on cellular signaling is central to molecular evolution. These questions are commonly examined using computational and statistical approaches. More recently, ancestral sequence resurrection (ASR) has combined inference of ancient protein sequences with experimental functional characterization. However, the reliance of these studies on low-throughput experiments limits them to examining a relatively small number of hypotheses. Currently, we have little direct, unbiased information about how protein function may change across large phylogenies. Here we develop an approach integrating large-scale ancestral sequence reconstruction with structural modeling, molecular dynamics, function prediction and experimental validation to characterize molecular-functional shifts and their structural bases across protein families with 1000s of sequences. We apply this approach to study two families of RNA-binding proteins spanning animal and plant lineages. We identify discrete shifts in protein-ligand affinities and long-term changes in function occurring across multiple nodes. Our results suggest that changes in protein function may not always be associated with gene duplication or major speciation events. The identification of functional 'flip flopping' - repeated transitions among a small number of functional states through different structural mechanisms - also supports the view that protein function may be highly evolutionarily labile. We characterize co-evolution between RNA binding proteins and their signaling partners, suggesting that co-evolutionary processes may be common, even when large shifts in molecular function are rare. We suggest that an 'unbiased' view of protein functional evolution may reveal new information about how protein families evolve when we aren't looking.

Yuri L. Lyubchenko

University of Nebraska Medical Center, USA

Title: Nanoscale structure and dynamics of centromere nucleosomes
Biography:

Yuri L. Lyubchenko is Professor of Pharmaceutical Sciences at the University of Nebraska Medical Center, Omaha, NE, USA. His research focuses on understanding fundamental mechanisms underlying health and disease, which are key to developing new and more effective diagnostics and medications. This primarily basic research allows him not only identify new drug targets for small molecule drugs, it also leads to development of the nanotools and methods to discover novel approaches for diagnostic, treatment and disease prevention and to more rapidly determine their efficacy at the molecular level.

Abstract:

Statement of the Problem: Chromatin integrity is crucial for normal cell development. The cell division process is accompanied by the segregation of replicated chromosome, and chromatin centromeres, specialized segments of chromosomes provide the accuracy of the chromosomal segregation. If the centromere becomes damaged or removed, chromosomes segregate randomly disrupting the cell division process. The centromeres are specifically recognized by kinetochores suggesting that centromeres contain specific structural characteristics. However, these structural details and the mechanism underlying their highly specific recognition remain uncertain. Methodology & Theoretical Orientation: In this study, we performed direct imaging of CENP-A nucleosome core particles by time-lapse high-speed atomic force microscopy (AFM), enabling us to directly visualize the dynamics of CENP-A nucleosomes. Nucleosomes used for evaluation of DNA wrapping around the histone core were assembled on a DNA substrate containing a centrally positioned 601 motif. Findings: A broadly dynamic behavior of the DNA flanks was first revealed by analysis of AFM images acquired in ambient conditions. Time-lapse imaging further identified the distinctive pathways unique to CENP-A-nucleosome dynamics that are not shared by H3. The spontaneous unwrapping of DNA flanks can be accompanied by the reversible and dynamic formation of loops with sizes equivalent to a single wrap of DNA. Translocation of CENP-A nucleosomes was observed, with the formation of internal DNA loops along the nucleosome. This process was reversible, settling the core back to its starting position. Additionally, the transfer of the histone core from one DNA substrate to another was visualized, as well as distinctive splitting into sub-nucleosomal particles that was also reversible. Conclusion & Significance: Altogether, our data suggest that unlike H3, CENP-A is very dynamic, permitting its nucleosome to distort freely and reversibly, which in turn allows a longer term stability, which may play a critical role in centromere integrity during mitosis and replication.   

Biography:

Andrei A. Korostelev is passionate about mechanisms of translation regulation. He received Ph.D. in Michael S. Chapman laboratory at Florida State University in 2003 and performed postdoctoral studies with Harry F. Noller in 2004-2010. The Korostelev laboratory at the RNA Therapeutics Institute uses recent advances in biochemical and structural methods to elucidate detailed mechanisms that govern translation and regulation of translation under stress conditions or during disease. Recent work revealed high-resolution “frames” of the motions that the translational machinery undergoes during bacterial stress responses (including the stringent response) and viral infection, as summarized on the laboratory web site: http://labs.umassmed.edu/korostelevlab/research.htm

Abstract:

Virus propagation depends on efficient synthesis of viral proteins by the host translational machinery. Internal ribosome entry sites (IRESs) of viral mRNAs mediate cap-independent initiation. Intergenic-region (IGR) IRESs of Dicistroviridae family, which includes the Taura syndrome virus (TSV) and Cricket paralysis virus (CrPV), use the most streamlined mechanism of initiation, independent of initiation factors and initiator tRNA. A tRNA-mRNA like pseudoknot of IGR IRESs binds the ribosomal A (aminoacyl-tRNA) site of the 80S ribosome (Fernandez et al., 2014; Koh et al., 2014). The pseudoknot has translocate to the P site to allow binding of the first tRNA and initiate translation.

Using electron cryo-microscopy of a single specimen, we resolved five ribosome structures formed with the Taura syndrome virus IRES and translocase eEF2•GTP bound with sordarin. The structures suggest a trajectory of IRES translocation, required for translation initiation, and provide an unprecedented view of eEF2 dynamics (animation).

The IRES rearranges from extended to bent to extended conformations. This inchworm-like movement is coupled with ribosomal inter-subunit rotation and 40S head swivel. eEF2, attached to the 60S subunit, slides along the rotating 40S subunit to enter the A site. Its diphthamide-bearing tip at domain IV separates the tRNA-mRNA-like pseudoknot I (PKI) of the IRES from the decoding center. This unlocks 40S domains, facilitating head swivel and biasing IRES translocation via hitherto-elusive intermediates with PKI captured between the A and P sites. 

  • Track 4: Signalling Biology | Track 5: Molecular Modelling and Dynamics | Track 6: Drug Designing
Location:

Session Introduction

Arfaxad Reyes Alcaraz

Korea University College of Medicine, South Korea

Title: Structural Conformational changes report biased agonism: The case of Galanin receptors
Biography:

Arfaxad Reyes has his expertise in structure and stability of G-protein coupled receptors and passion for improving and creating new drug discovery platforms that greatly contribute in the development of more selective drugs with minor side effects. His studies about biased agonism in Galanin receptors help to understand the relationship between conformational structure of the receptor and its corresponding physiological effect induced by a specific ligand. Recently he and co-workers were able to develop a highly selective agonist for Galanin receptor 2 with anxiolytic effect in vivo (Arfaxad Reyes-Alcaraz et al Sci. Rep 2016) which was the base to discover how different ligand structures induce different conformations on the structure of Galanin receptors. This work greatly contribute to understand the relationship between structure and function of Galanin receptors.

Abstract:

Statement of the Problem: G protein coupled receptors (GPCRs) also known as seven-transmembrane receptors and are the largest family of cell-surface receptors that communicate extracellular stimuli to the cell interior (1). It is now accepted that chemically distinct ligands bind to the same GPCR and can stabilize the receptor in multiple active conformations, which results in differential activation of cell signaling pathways and, eventually, in different physiologic outcomes a phenomenon known as biased agonism (2). Biased agonism can be exploited to design drugs that selectively activate signaling pathways, leading to the desired physiologic effects while on target side effects elicited by activation of other signaling pathways via the same receptor subtype (3). Methodology & Theoretical Orientation: The aim of this study was to stablish a relationship between conformational changes in Galanin receptors and their signaling properties in living cells, for that purpose we develop a structural complementation assay based on NanoBit technology and a series of conformational fluorescein arsenical hairpin (FIASH) bioluminescence resonance energy transfer (BRET) biosensors to monitor structural changes β-arrestin 2 induced by the binding with each Galanin receptor. Findings: Here we show that Galanin receptors impose different conformational signatures in β-arrestin, moreover structurally different ligands activating the same receptor imposed different conformations in β-arrestin 2 producing biased signaling. Conclusion & Significance: Our data provide definite evidence that a receptor activated by structurally different ligands can adopt multiple active conformations. Moreover, this finding also demonstrates that functionally specific structural Galanin receptor conformations can indeed be translated to downstream effectors producing a different physiological response. 

Biography:

Vesa P. Hytönen is a head of the Protein Dynamics research group in BioMediTech at the University of Tampere, Tampere, Finland. After graduating as a PhD from the University of Jyväskylä, Jyväskylä, Finland at 2005, he conducted postdoctoral training at ETH Zurich, Zürich, Switzerland 2005–2007. He then continued as a postdoctoral researcher at the University of Tampere and established independent research group at 2010. He is currently working as Associate Professor at the University of Tampere. His research interests are mechanobiology, protein engineering and vaccine research and he has authored more than 100 scientific articles.

Abstract:

Talin is a central adhesion protein linking β-integrin cytosolic domains to actin fibers. It participates in the transmission of mechanical signals between extracellular matrix and cell cytoskeleton. Talin rod domain consists of a series of mechanically vulnerable α-helical subdomains containing binding sites for other adhesion proteins such as vinculin, actin and RIAM. Force induced unfolding of these rod subdomains has been proposed to act as a cellular mechanosensor, but so far evidence linking their mechanical stability and cellular response has been lacking. We show that stepwise mechanical destabilization of talin rod subdomain increases talin and vinculin accumulation into cell-matrix adhesions and decreases cell migration rate. In addition, mechanical destabilization of talin subdomain was found to decrease cellular traction force generation and to promote the formation of adhesions on fibronectin over vitronectin. Experiments with truncated talin forms confirmed the mechanosensory role of the talin subdomain and excluded the possibility that the observed effects are caused solely by the release of talin autoinhibition. We demonstrate that by modulating the mechanical stability of an individual talin rod subdomain, it is possible to affect traction force generation, ECM sensing and consequently highly coordinated processes such as cell migration. Our results suggest that talin acts as a mechanosensor and is responsible for controlling the cellular processes dependent on mechanical signals and cellular mechanosensing.

Joachim Krebs

Max Planck Institute for Biophysical Chemistry, Germany

Title: Calcium, Calmodulin and the Plasma Membrane Calcium Pump
Biography:

Joachim Krebs has been working in the field of calcium-binding and calcium-transporting proteins for many years. After receiving his PhD he spent 2 years as a postdoctoral fellow in the Lab of Prof. RJP Williams at Oxford, UK. In 1977 he accepted a staff position at the Institute of Biochemistry at the Swiss Federal Institute of Technology (ETH) in Zürich, Switzerland. He was lecturing different courses in biochemistry and biophysics, and was leading a Lab working on the structure-function relationship of calcium-binding and calcium-transporting proteins. After retirement from the ETH he continued his research as a consultant of the Lab of Prof. Christian Griesinger at the MPI in Göttingen, Germany. He has authored, co-authored and edited numerous articles in international journals. Recently he edited a book on “Calcium: A matter of life or death” published in 2007. He is at the Editorial Board of BBA Molecular Cell Research and Archives of Biochemistry and Biophysics.

Abstract:

Calcium is the third most abundant metal in nature and a versatile carrier of many signals within and outside the cell. Due to its peculiar coordination chemistry calcium is highly flexible as a ligand which enables it to regulate many important aspects of cellular activity. Calcium can fulfill its many different functions insite and out of the cell due to an integrated network of calcium channels, exchangers and pumps. In this presentation I will give an overview on our studies of calcium binding proteins, their interaction with protein targets resulting in specific modulations  of protein-protein interactions. This will be demonstrated by the interaction of the calcium binding protein calmodulin with one of its targets, the plasma membrane calcium pump, an important regulator of calcium homeostasis of the cell.

Biography:

Igor Barsukov has expertise in structural biology, primarily using NMR spectroscopy and X-ray crystallography. The main focus of his research has been on the structure and function of integrin-mediated cell-matrix adhesions, where he directed full structural analysis of the key adhesion proteins talin, leading to the currently widely used model of stretch-dependent talin activation. He is currently extending the model of talin functionality to include competitive, talin based, interaction networks. Recently he identified interactions between neuronal scaffold proteins Shank3 and Ras-family GTPase that regulate integrin activity and have implications for control of synaptic plasticity.

Abstract:

Statement of the Problem: Cell-matrix adhesion requires assembly of large multi-protein complexes linked to the cyto-domains on the integrin adhesion receptors. These complexes dynamically change in response to the adhesion forces and environmental signals. Mechano-sensitive adaptor protein talin couples the force and adhesion signaling by acting as a hub for numerous, often competitive, interactions. The molecular mechanism that controls the co-ordination of the interactions in time and space is currently not understood. The aim of this study is to define the interactions between talin and Rho-GAP Deleted in Liver Cancer 1 (DLC1) that regulates adhesion forces. Methodology: Structure of the talin/DLC1 complex was solved by X-ray crystallography and the interactions between the proteins analysed by NMR spectroscopy. Fluorescent imaging was used to define the interactions within the adhesion complexes and cancer cell lines were employed to characterise the effect of the interaction on the biological activity. Findings: We defined the atomic details of the talin/DLC1 interactions and used this information to identify signalling protein paxillin as a talin ligand. Based on the structural information we designed a range of talin mutants that modulate the interactions and demonstrated that the mutations reduce DLC1 signalling in adhesion. Conclusion & Significance: Talin recognises LD motifs in DLC1 and paxillin through a set of well-defined charge interactions. These interactions are similar to other LD motif interactions previously identified in signalling pathways. These interactions are also similar to the interactions between talin, RIAM and vinculin that were previously not assigned to the LD motif family. Together, the relatively weak LD motif interactions within the adhesion complex create a protein network that could dynamically respond to the adhesion signals.

Biography:

Shuanghong Huo received her Ph.D. in Computational Chemistry from Boston University. She had postdoctoral training at UC-San Francisco. She is a Professor of Chemistry and Biochemistry at Clark University, Worcester, USA. Her research interest is protein folding, misfolding, and aggregation. Recently her group is developing dimensionality reduction methods and graph representations of protein free energy landscapes. 

Abstract:

In the study of protein thermodynamics and kinetics it is of paramount importance to characterize protein free energy landscapes. Dimensionality reduction is a valuable tool to complete the task. We have evaluated several methods of dimensionality reduction, including linear and non-linear methods, such as principal component analysis, Isomap, locally linear embedding, and diffusion maps. A series of criteria was used to assess different aspects of the embedding qualities. Our results have shown that there is no clear winner in all aspects of the evaluation and each dimensionality-reduction method has its limitations in a certain aspect. The linear method, principal component analysis, is not worse than the nonlinear ones in some aspects for our peptide system. We have also developed a mathematical formulation to demonstrate that an explicit Euclidean-based representation of protein conformation space and the local distance metric associated to it improve the quality of dimensionality reduction. For a certain sense, clustering protein conformations into macro-clusters to build a Markov state model is also an approach of dimensionality. We have tested inherent structure and geometric metric for state space discretization and demonstrated that the macro-clusters based on inherent structure give a meaningful state space discretization in terms of conformational features and kinetics.

Biography:

Head, Laboratory of Biological Physics, Institute of Physics, Polish Academy of Sciences in Warsaw, Poland. Education - M.S., Department of Physics, University of Warsaw, 1973; Ph. D., Department of Physics, University of Pittsburgh, 1977; D.Sc., Department of Physics, University of Warsaw, 1984. Professorial title, 1994. Fields of interest – condensed matter theory (spin waves, spin glasses, porous media, growth processes, atomic friction, river networks, nanofluidics, self-organized nanostructures) and biological physics (large conformational changes of biomolecules within coarse-grained models, especially as induced by stretching,  proteins with knots and slipknots, protein folding, dynamics of virus capsids and other multi-proteinic structures such as a cellulosome, interaction of proteins with solids, proteins at air-water interface, modeling of proteasomes,  inference of genetic networks from the microarray data). Co-author of textbook “Theory of Quanta”, Oxford University Press 1992. 250 research papers.

Abstract:

We review the physics of  processes involving large conformational transformations in knotted proteins in bulk water and then consider folding in ribosomes and unfolding in proteasomes.  Formation of a knot is demonstrated to be facilitated by the nascent conditions at the ribosome. Knots in proteins have been proposed to resist proteasomal degradation. Ample evidence associates proteasomal degradation with neurodegeneration. One interesting possibility is that indeed knotted conformers stall this machinery leading to toxicity. However, although the proteasome is known to unfold mechanically its substrates, at present there are no experimental methods to emulate this particular traction geometry. Here, we consider several dynamical models of the proteasome in which the complex is represented by an effective potential with an added pulling force. This force is meant to induce translocation of a protein or a polypeptide into the catalytic chamber. The force is either constant or applied periodically. The translocated proteins are modelled in a coarse-grained fashion. We do comparative analysis of several knotted globular proteins and the transiently knotted polyglutamine tracts of length 60 alone and fused in exon 1 of the huntingtin protein. Huntingtin is associated with Huntington disease, a well-known genetically-determined neurodegenerative disease. We show that the presence of a knot hinders and sometimes  even jams translocation. We demonstrate that the probability to do so depends on the protein, the model of the proteasome, the magnitude of the pulling force, and the choice of the pulled terminus. In any case, the net effect would be a hindrance in the proteasomal degradation process in the cell. This would then yield toxicity via two different mechanisms: one through toxic monomers compromising degradation and another by the formation of toxic oligomers

Biography:

Christina Scharnagl has her expertise in molecular dynamic simulations of membrane proteins. Her work focuses on biophysical principles of the interdependence of transmembrane helix dynamics, helix-helix binding, and helix-lipid interactions. In silico modelling and advanced computational analysis are closely connected to experimental work in research collaborations in order to interpret and guide the experiments and to validate the simulations. The aim of the joint efforts is to understand the impact of these phenomena on multiple biological processes, such as membrane fusion and intramembrane proteolysis. 

Abstract:

Integral membrane proteins facilitate communication between the inside of the cell and its exterior. Their transmembrane domains (TMDs) support a diversity of biological functions and exhibit sequence-dependent conformational dynamics on multiple size and time scales. Membrane proteins are notoriously difficult to study by experimental methods. Molecular dynamics (MD) simulations provide a powerful tool of high spatial and temporal resolution that effectively complements experimental methods. Here we focus on the conformational dynamics of the TMD of the amyloid precursor protein (APP). APP is enzymatically hydrolyzed within its TMD by g-secretase (GSEC), forming toxic Ab peptides regarded as molecular cause of Alzheimer's disease (AD). Besides APP, GSEC cleaves ~100 single-span membrane proteins within their TMDs, however without obvious consensus sequence. Finding the link between the molecular architecture of the substrate TMDs and cleavage is, therefore, of upmost importance. Because unfolding is obvious to expose the scissile bond, it seems plausible that the TMD itself is optimized for local helix unwinding. However, this view was challenged by our experiments and MD simulations. Our results suggest an alternative model where reaching a cleavage-competent state involves multiple conformational transitions of the substrate/enzyme complex where global conformational plasticity of the substrate TMD is a key determinant. In a first step, we compare the conformational flexibility of a large number of substrate and non-substrate TMDs, as well as TMDs carrying missense mutations related to early onset AD. Knowing the key-dynamical motifs will help to identify new substrates and to elucidate the physiological functions of the protease in the brain and other organs. This work is part of a collaborative research program (https://www.i-proteolysis.de/). 

Biography:

Sadeq Vallian finished his PhD degree at King’s College London, London in 1996. Then he did his post-doctoral fellowship in Laboratory Medicine at M.D. Anderson Cancer Center, Texas University, Texas in 1999. He then moved to University of Isfahan, Isfahan, IR Iran, and started his independent research work as an assistant professor in 2000. Now he is a full-time professor in the department of biology, at University of Isfahan. His research was mainly focused on human genome variation and genetic diseases and cancer. In the last two years, two projects have been running in his lab, i) the effects of genomic variation and non-synonymous polymorphisms in drug/target interactions in cancer chemotherapy especially those affecting Topoisomerase II and its targeting drugs; ii) Genomic polymorphism and non-coding RNA, their expression and target interactions. 

Abstract:

Topoisomerases are the key enzymes involved in resolving the winding of DNA during DNA replication, transcription, recombination, and chromatin remodeling. Because of the essential roles of these enzymes in the maintenance of genome stability and cell function, topoisomerases were the important targets for cancer chemotherapy drugs. To date, several topoisomerase inhibitors have been introduced and applied as drugs in the treatment of cancer. Topoisomerase II α (Top2-α), a subclass of topoisomerase II enzymes, functions as an important target for several anticancer drugs. It seems that genomic variations such as non-synonymous polymorphisms (nsSNP) could affect the enzyme/drug interactions. Our main focus has been the study of the mechanisms by which non-synonymous genomic variations in Top2-α could affect its interaction with anticancer drugs such as Amsacrine and Mitoxantrone as important inhibitors of the enzyme. In the first step we identified the main residues involved in the interaction between the enzyme and the drugs. In <5°A space around the drug, the main residues were Lys489, Asn504, Glu506, Arg487 and Ile490 for Mitoxantrone, and for Amsacrine, the interaction was through Glu506, Leu491, Arg487, Gly488, Lys489, Ile490 and Asn504 residues. Next, the effects of non-synonymous polymorphisms (nsSNPs) were examined on the interaction of Top2-α with the drugs. The results showed that variants K529E, R568H, R568G, T530M and R487K, Y481C could affect the inhibition by Amsacrine and Mitoxantrone, respectively. This suggested that nsSNPs could clearly affect the inhibition of Top2-α causing possible drug-resistant. These results could facilitate the prediction and development of drugs for specific Top2-α inhibition, making the cancer chemotherapy more effective.