Scientific Program

Conference Series Ltd invites all the participants across the globe to attend 10th International Congress on Structural Biology Helsinki, Finland.

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Scientific Program Day 1
Scientific Program Day 2

Day 1 :

Keynote Forum

Annette G Beck-Sickinger

Leipzig University, Germany

Keynote: Structural insights in the binding mode of neuropeptide Y at G protein coupled receptors and consequences for drug development
Conference Series Structural Biology Meet 2018 International Conference Keynote Speaker Annette G Beck-Sickinger photo
Biography:

Annette G. Beck-Sickinger studied chemistry and biology at the University of Tubingen (Germany) and received her Ph. D. in organic chemistry.  She was post-doc with E. Carafoli (Laboratory of Biochemistry, ETH Zurich) and appointed as assistant professor of Pharmaceutical Biochemistry at ETH Zurich. Since October 1999, she is full professor of Biochemistry and Bioorganic Chemistry at the University of Leipzig. She spent a sabbatical at Vanderbilt University (Nashville, TN) as visiting professor. Annette Beck-Sickinger was a member of the Board of the German Chemical Society (Gesellschaft Deutscher Chemiker, 2004-2012; Vice-President 2006-2008) and of the DFG panel "Biochemistry“ (2004-2012). Since 2017 she is member of the Board of the German Society for Biochemistry and Molecular Biology (gbm) and Vice-President. She has been awarded protein-coupledmany prices including the Leonidas Zervas Award of the European Peptide Society, the gold medal of the Max-Bergmann-Kreis (2009), the Leipzig Science Award (2016) and the Albrecht Kossel Award of the GDCh (2018). She was honoured with the membership of the Saxonian Academy of Science in 2009 and in 2012, she became an elected member of the German National Academy of Sciences Leopoldina. In 2017, she was awarded cross-linkingthe Saxonian Order of Merit. Her major reseach fields are structure-activity-relationships of peptide hormones and G protein coupled receptors and protein modification to study function and interaction. A tight connection of chemical methods, bioorganic synthesis and molecular biology tools, including cloning, receptor mutagenesis, protein expression and cell biochemistry is applied. Her interests include identification of novel targets and novel therapeutic concepts and novel approaches to modify proteins and concepts for improved enzyme catalysis and biomaterials.

 

Abstract:

Peptides hormones play an important role in the regulation of manifold activities in the body. Many of them transmit their activity through G-Protein Coupled Receptors (GPCR), which are among the most promising drug targets nowadays. Accordingly, elucidating the binding mode of ligands is essential. Whereas several small molecule systems have been well characterized, ligand binding of large and flexible peptides is still more challenging. In addition to ligand binding and receptor activation, indirect mechanisms have been shown to play a role for drugs addressing GPCRs. This includes desensitization, internalization and accordingly their potential use as drug shuttles, e.g. in tumor targeting. Accordingly, in addition to ligand binding, internalization has to be addressed and to be studied, including arrestin recruitment. Accordingly, ligand binding, structural dynamics, and internalization have to be addressed and to be studied to address G protein-coupled receptors for drug development. The neuropeptide Y/pancreatic polypeptide family contains 36 amino acid peptides that bind in human to four different so-called Y-receptors. By a combination of X-ray analysis, NMR, molecular modeling and cross-linking combined with mass spectrometry, we could recently identify the distinct binding modes of NPY to the Y1- and the Y2 receptors. We could further demonstrate that chemical modification of the ligand, including fluorescence labeling, lipidization, and PEGylation significantly modifies the trafficking of the ligand. By labeling of the receptor with a novel template-assisted ligation strategy, we can follow ligand/receptor complexes in living cells. Furthermore, we identified a different mode of arrestin binding and recruitment. Neuropeptide Y1 and Y2 receptors have been shown to play a relevant role in different tumors. In breast cancer we demonstrated that human Y1 receptors are addressable by peptide conjugates using 99mTc or 18F PET-tracers. We now designed Y1 receptor selective peptides linked to different toxophors. Furthermore, we characterized the mechanism of direct and peptide-mediated uptake of tubulysin-related toxins. In the field of tumor therapy, peptide-drug conjugates are already well accepted. However, the concept of receptor-mediated internalization and subsequent tissue specific intracellular application is not limited to the selective addressing of tumors. This may open up a new field of targeted therapy by mid-sized drugs.
 

  • 3D Structure Determination| Computational Approaches in Structural Biology| Hybrid Approaches for Structure Prediction| Structural Bioinformatics and Proteomics
Location: Alto
Speaker

Chair

Annette G Beck-Sickinger

Leipzig University

Session Introduction

Ozge Sensoy

Istanbul Medipol University, Turkey

Title: 11:30 - 12:10
Speaker
Biography:

Ozge Sensoy has her research studies focused on understanding molecular mechanisms of biologically important systems and also providing mechanistic insight at the molecular level. She has been working with GPCRs and their interacting partners which are responsible for cellular signaling. She has been awarded an international COST grant which is based on developing heterobivalent molecules capable of binding more than one target for treatment of symptoms of Parkinson’s disease.
 

Abstract:

Parkinson’s disease is caused by disruption of cells which provide dopamine to the striatum in the brain. Therefore, the main approach made for treatment of the disease has based on increasing dopaminergic signaling by using dopamine agonists, preventing dopamine breakdown via monoamine oxidase enzymes or supplying additional dopamine in the form of L-dopa. Even though L-dopa is known as the most effective drug so far, its efficacy decreases with time due to the use of high dosage of the drug. Moreover, it also causes motor complications such as motor fluctuations and dyskinesia. In this multidisciplinary project, we have aimed for developing hetero-bivalent ligands that target A2AR (Adenosine-2A-receptor)-D2R (Dopamine-2receptor) hetero-tetramer, which has been shown to be the dominant stoichiometry of A2AR-D2R. Firstly, we design and dock hetero-bivalent ligands to the receptors and investigate the molecular mechanism of allosteric interactions within the heterotetramer by means of accelerated molecular dynamics simulations. Subsequently, successful drug candidates are synthesized and tested in vitro for their activities. More importantly, the drug candidates are also tested by in silico and in vitro models for their permeation against blood-brain barrier. So far, hetero-bivalent ligands have been only used as molecular tools for detecting the existence of the receptor dimers. On the other hand, the current study will provide a chance to test the capability of hetero-bivalent ligands for being used as therapeutic molecules. In this way, we expect to develop more effective therapeutic molecules to alleviate the symptoms of Parkinson’s disease hence increasing the quality of patient’s life

Ozge Sensoy

Istanbul Medipol University

Title: 11:30 - 12:10
Biography:

Ozge Sensoy has her research studies focused on understanding molecular mechanisms of biologically important systems and also providing mechanistic insight at the molecular level. She has been working with GPCRs and their interacting partners which are responsible for cellular signaling. She has been awarded an international COST grant which is based on developing heterobivalent molecules capable of binding more than one target for treatment of symptoms of Parkinson’s disease.
 

Abstract:

Parkinson’s disease is caused by disruption of cells which provide dopamine to the striatum in the brain. Therefore, the main approach made for treatment of the disease has based on increasing dopaminergic signaling by using dopamine agonists, preventing dopamine breakdown via monoamine oxidase enzymes or supplying additional dopamine in the form of L-dopa. Even though L-dopa is known as the most effective drug so far, its efficacy decreases with time due to the use of high dosage of the drug. Moreover, it also causes motor complications such as motor fluctuations and dyskinesia. In this multidisciplinary project, we have aimed for developing hetero-bivalent ligands that target A2AR (Adenosine-2A-receptor)-D2R (Dopamine-2receptor) hetero-tetramer, which has been shown to be the dominant stoichiometry of A2AR-D2R. Firstly, we design and dock hetero-bivalent ligands to the receptors and investigate the molecular mechanism of allosteric interactions within the heterotetramer by means of accelerated molecular dynamics simulations. Subsequently, successful drug candidates are synthesized and tested in vitro for their activities. More importantly, the drug candidates are also tested by in silico and in vitro models for their permeation against blood-brain barrier. So far, hetero-bivalent ligands have been only used as molecular tools for detecting the existence of the receptor dimers. On the other hand, the current study will provide a chance to test the capability of hetero-bivalent ligands for being used as therapeutic molecules. In this way, we expect to develop more effective therapeutic molecules to alleviate the symptoms of Parkinson’s disease hence increasing the quality of patient’s life

Meriem El Ghachi

University of Liege, Belgium

Title: 12:10 - 12:50
Speaker
Biography:

Meriem El Ghachi has completed her PhD from Paris University of Paris-Sud, France. He is a Post-doctorate at University of Liege, Belgium. She has published 17 papers in reputed journals.

Abstract:

Undecaprenyl-phosphate C55-P is a key lipid carrier of glycan intermediate required for the synthesis of a variety of cell wall polymers such as Peptidoglycan (PG), Lipopolysaccharides (LPS) O-antigen wall teichoic acids, capsular polysaccharide, common enterobacterial antigen, membrane-derivative oligosaccharides and exopolysaccharides. In bacteria, during peptidoglycan synthesis, the phospho-N-acetylmuramoyl (-pentapeptide) -N-acetyl glucosamine is the essential motif carried by the C55-P. The resulting lipid, C55-PP-MurNAc-(pentatpetide)-GlucNAc (lipid II), is translocated towards the periplasmic side by several putative flippases. The MurNAc-(pentatpetide)-GlucNAc is added to the elongating chains of PG and C55- P is released as C55- PP. This precursor is also provided by the de novo synthesis in the cytosol that is catalyzed by a Cis-Prenyl Pyrophosphate Synthase, UPPS, which successively adds eight isoprene units from C5-PP on farnesyl pyrophosphate. Two families of phosphatases can perform the subsequent dephosphorylation of C55-PP into C55-P, common to the in vitro synthesis and carrier lipid recycling. In E. coli, 1 BacA and 3 phosphatidic acid phosphatases 2 (PgpB, YbjG and LpxT), active on C55-PP have been identified. PgpB being also involved in the phosphatidyl glycerol metabolism and LpxT transferring the phosphate from C55-PP to lipid A. Whereas Bacillus subtilis has three C55-PP phosphatases 1 BacA (YubB) and 2 phosphatidic acid phosphatases 2 (YwoA and YodM). We obtained the structure of BacA using lipidic cubic phase method. The crystal structure at 2.6 A revealed an unexpected fold according to the previous biochemical studies. Moreover, we solved the structure of bsPgpB (yodM) in the presence and absence of its favorite substrate, phosphatidyl glycerol. 

Ghazaleh Gharib

Sabanci University Nanotechnology Research and Application Center, Turkey

Title: Cloning, sequencing, purification and characterization of highly thermostable aspartate aminotransferase from Geobacillus thermopakistaniensis
Speaker
Biography:

Ghazaleh Gharib received her Bachelor’s degree in General Biology from Ferdowsi University of Mashhad, Iran in 2004. She completed her MSc in Biotechnology from Institute of Biochemistry and Biotechnology University of the Punjab in 2008 and then completed her PhD in 2016 from School of Biological Sciences, University of the Punjab under HEC scholarship. Her work is majorly focused on study of hyperthermophiles and archaeal enzymes. She is currently pursuing her Postdoctoral research studies on a project approved by TUBITAK International fellowship in Sabanci University Nanotechnology Research and Application Center, Istanbul, Turkey.
 

Abstract:

Aspartate aminotransferase gene consists of 1,182 bp nucleotide encodes for 393 amino acids was sequenced, cloned, expressed and purified to homogeneity. Enzyme exhibited maximum activity at pH 7 at 65 ºC. Mass spectrometry determined molecular mass of 42,562 Da and gel filtration indicated the protein exist as a dimeric form. Thermo stability experiment showed 100% stability of a protein at 65 ºC for 16 hours and half-life of 15 mins at 75 ºC. The thermal denaturation studies by CD spectroscopy showed no significant change in ellipticity of the helical structure of the protein below 70 ºC. Km and Vmax values towards aspartate were 1.61 mM and 97 µmol min-1 mg-1 and towards α-ketoglutarate were 2.5 mM and 50 µmol min-1 mg-1, respectively. Substrate specificity experiment indicated maximum activity with aspartate and its respective keto acid (α-ketoglutarate ) while exhibits 27% of activity with Tyr and 16% of its activity with Pro and Cys and no activity with Glu how ever in reverse reaction of Glutamate with its keto acid, oxaloacetate, 70% of activity was observed. Pyridoxal phosphate quantification exhibited 0.1 mole of PLP per mole of enzyme. Amino acid analysis showed high contents of charged and especially acidic residues of Aspartate and glutamate in structure of ASTSBS enzyme. Homology modeling determined the Dimeric structure of ASTSBS which contained high number of proline on a surface of each sub-units as compare to its mesophilic counter parts that is a reason for stability of a protein at high temperature.
 

Sushil Tripathi

University of Helsinki, Finland

Title: Systematic transcriptomic profiling defines critical LKB1substrates in gastrointestinal tumorigenesis
Speaker
Biography:

Sushil Tripathi is a postdoctoral researcher in University of Helsinki. He had completed his PhD from Norwegian University of Science and Technology, Norway. 

Abstract:

The mechanism by which loss-of-function mutations of the LKB1 kinase lead to Peutz- Jeghers Syndrome (PJS) polyposis is unknown. Based on PJS models the characteristic hamartomatous polyps are driven by clonally expanding stromal myofibroblasts providing a starting point for molecular studies. LKB1 phosphorylates and thereby activates at least 14 related family kinases including AMPKa1, AMPKa2, NUAK1, NUAK2, SIK1, SIK2, SIK3, SNRK, BRSK1, BRSK2, MARK1, MARK2, MARK3 and MARK4. Genetically, LKB1 has been implicated as a regulator of metabolism, polarity, cell growth and migration and the LKB1 substrate kinases are partly linked to these cellular events. However, it is unknown, which of the LKB1 substrates are critical for LKB1-mediated tumor suppression. To investigate this, we established an in vitro system to study molecular changes following shRNA-mediated depletion of Lkb1 substrate kinases in cultured fibroblasts with myofibroblast characteristics. In this system, loss of Lkb1 leads to transcriptomic changes (Lkb1-signature) with a highly significant overlap with changes observed in Lkb1-deficient tumors in vivo, thus allowing for further defining a 134 gene Lkb1-myofibroblast signature possibly involved in driving tumorigenesis. Gene Set Enrichment Analysis (GSEA) of Lkb1- myofibroblast signature genes indicates that the Lkb1-deficiency is associated with signatures of Cancer-Associated Fibroblasts (CAFs) and extracellular matrix organization. Depletion of 14 Lkb1 substrate kinases lead to variable overlaps with Lkb1-signature genes and cumulatively the overlap is 75% supporting their importance in mediating Lkb1 functions. We have validated some these substrates and their commonly regulated genes for their involvement as potential mediators of tumorigenesis.
 

Maryam Alobathani

University of Sharjah, UAE

Title: Prevalence of Non-syndromic hearing loss in UAE due to genetic mutation in Mitochondrial 12S rRNA
Speaker
Biography:

Maryam Alobathani has completed her Bachelor studies in Biotechnology by age 22 from University of Sharjah and know she’s working as a laboratory supervisor and completing her Master studies at the same university.

 

Abstract:

Deafness is a disease of inability to hear referred as hear impairment or anacusis for those with low or no hearing. It’s important at all levels of health care due to their significant burden on affected individuals and societies. Hearing loss can be classified as partial or completely inability to hear. Deafness can occur due to different mutations in different human genome such as Connexin26 the most common deafness gene, CDH23, TMC1 and in mitochondria. mitochondrial organelle is ubiquitous in eukaryotic as intracellular double membrane structure. The main function of mitochondria is to synthesis ATP by OXPHS and plays important roles in cell death and oxidative stress control. Mutation in mitochondrial DNA cause OXPHS defects that leads to many diseases one of them is hearing loss as syndromic and non-syndromic deafness. Therefore, we aim in this research to identify the genetic causes of NSAHL underlying wide spectrum of congenital conditions in UAE population. This study focused on a non-syndromic deafness caused by mutation on a mitochondrial MTRNR1 gene that encodes 12SrRNA. 40 Samples were collected from unrelated NSHL UAE family members and screened using biological tools such as PCR, Genomic sequencing machines and Bioinformatics tools. Expert geneticist recruiting and clinically assessing the participant patients involved in this study.The main approaches proposed are: (i) homozygosity maps were used to localize the homozygous regions in each particular family, (ii) next generation sequencing platform used to sequence the whole-exome of an affected individuals to explore all variants including the pathogenic mutations, finally (iii) pathogenicity of the outcoming variant results validated or ruled out by performing functional assays. The study indicates two out of the total 40 samples of unrelated UAE families with deafness due to mitochondrial 12S rRNA mutation m.669 T>C and m.827A>G, as well some known polymorphisms. This study is part of other researches done on a larger scale on similar area of interest that will reach to figure out new mutation related to mtDNA 12S rRNA variations. This finding can also help in future genetic counselling, prenatal screening, and post-natal genetic diagnosis to prevent the prevalence of more NSHL patients.

  • Sequencing | Structural Molecular Biology | Structural Biology in Cancer Research| Signalling Biology
Location: Alto
Speaker

Chair

Annette G Beck-Sickinger

Leipzig University

Session Introduction

T V Koshlan

Peter the Great St. Petersburg Polytechnic University, Russia

Title: Identification of active sites interaction of different protein molecules in case of formation Nap1-Nap1, Mdm2-Mdm2 and P53-Mdm2

Time : 10:00 -11:00

Biography:

T V Koshlan is currently working as a Professor at Peter the Great St. Petersburg Polytechnic University, Institute of Applied Mathematics and Mechanics, Department of Higher Mathematics. He has completed his PhD in Physics and Mathematics with Mathematical modeling of the optical properties of multilayer biological systems and structures in their heterogeneous conjugation. He has habilitation at the State Polytechnic University of St. Petersburg, Russia. His research interests are diffraction theory, electrodynamics, physics of lasers, tissue optical methods of mathematical modeling in biological tissue optics and numerical method, biophysics.

Abstract:

In this report, two algorithms are developed, algorithm 1 and algorithm 2. Algorithm 1 was developed in order to search for the interaction of a polypeptide chain of a full-length protein with short active region. Algorithm 2 was developed to determine the most active sites of interaction between full-length proteins when dimers are formed in the direction from the N terminus to C terminus. Numerical calculations were made using proteins Mdm2, Nap1, P53. For modern proteomics, research and prediction of protein interactions are very important tasks, since they determine the function of proteins at levels from the cell to the whole organism. For proteins whose structure is known, the search for intermolecular interactions according to known data on the conformation of their tertiary structure reduces to the problem of searching for geometric complementarity of the sections of two interacting molecular surfaces and modeling their contacts, the so-called molecular docking. The task of molecular docking is the task of a conformational search algorithm, which reduces to a search for the conformational space of the formed biological complex due to the variation of the torsion angles of protein molecules. Modern conformational search algorithms in most cases find conformations that are generally close to the experimentally found structures in a relatively short time. However, there are factors that also have a significant impact on the success of the docking, which are often not taken into account in standard algorithms. One such factor is the conformational mobility of the target protein. The mobility range can be different beginning with a small adjustment of the side chains and ending with scale domain movements. These movements play an important role. At first glance, the most logical solution to this problem is to take into account the mobility of the protein in a docking program. Unfortunately, modern computational tools do not allow such modeling to be performed in an acceptable time frame since a protein molecule is very large, and allowing for mobility over all degrees of freedom can lead to a so-called combinatorial explosion (an astronomical increase in the number of possible variants). Only in some programs is there a limited mobility of protein binding sites (usually at the level of a small adaptation of conformations of the side chains of the active center residues). Another approach to this problem consists in docking the same protein in several different conformations and then selecting the best solutions from each docking run. The third approach is to find a universal structure of the target protein in which docking would produce fairly good results for different classes of ligands. In this case, the number of missed (but correct) solutions decreases, but the number of incorrect options also increases significantly. It should also be noted that most programs for the theoretical docking of proteins work according to the following principle: one protein is fixed in space, and the second is rotated around it in a variety of ways. At the same time, for each rotation configuration, estimates are made for the evaluation function. The evaluation function is based on surface complementarity (the mutual correspondence of complementary structures (macromolecules, radicals), determined by their chemical properties), electrostatic interactions, van der Waals repulsion and so on. The problem with this approach is that calculations throughout the configuration space require a lot of time, rarely leading to a single solution, which in turn does not allow us to speak of the uniqueness of the target protein and ligand interaction variant. So in the work while modeling by the methods of molecular dynamics, from 200 to 10 000 possible combinations of the formation of a protein complex with a ligand were found. Such a large number of modifications, along with the lack of a criterion for selecting the most probable variants of the bound structures of biological complexes (which would allow a radical reduction in their number) makes it very difficult to interpret the theoretical results obtained for practical use, namely, the finding of catalytic centers and a qualitative assessment of the dissociation constant of interacting substances. In contrast to the above computer simulation algorithms, mathematical algorithms have been developed in this chapter that allow determining the detection of proteins active regions and detecting the stability of different regions of protein complexes (linear docking) by analyzing the potential energy matrix of pairwise electrostatic interaction between different sites of the biological complex, such as the homodimer of the histone chaperone Nap1-Nap1, the heterodimer of the p53 Mdm2 proteins, and the homodimer Mdm2 Mdm2, which are responsible for the entry of a whole protein molecule into biochemical reactions.

Mohd Athar

Central University of Gujarat, India

Title: First protein drug target’s appraisal of lead-likeness descriptors to unfold the intervening chemical space
Speaker
Biography:

Mohd Athar is a Senior Research Fellow in Computational Chemistry Group at Central University of Gujarat. He holds the DST-INSPIRE Fellowship awarded by the Ministry of Science and Technology (DST) India. He has completed his Master’s degree in Applied Chemistry at BBA Central University Lucknow and Bachelor’s degree in Biotechnology from Hemwati Nandan Bahuguna Garhwal University. His major research is in the area of Medicinal Chemistry ranging from Drug Discovery, Combinatorial Chemistry, In silico Virtual Screening, Lead Optimization/Designing, Target Identification to its Validation.
 

Abstract:

A plethora of literature has been published for unveiling the problems associated with lead and drug likeness. However, despite of these advances in combinatorial chemistry, high throughput methods and virtual screening, plethora of clinical studies disquiets due to lead and drug-likeness attrition. For mitigation, the knowledge of physicochemical properties is really useful for guiding the design and selection of compounds from libraries dictated by certain rule of thumbs. However, robust biotechnological and instrumental innovations have created exponential increase in novel compounds and databases which compelled rethinking of evaluation procedures. Known descriptive molecular property filters proposed by Lipinski, Verber and Hann are not efficient enough to encompass long array of compounds and do not take into account the specificity of biological target. In this pursuit, we have tried to appraise eight molecular properties for two major classes of biological targets viz. membrane proteins and ion channels binding ligands. It has been proposed that the target based knowledge of descriptors can guide the selection of molecules to pick compounds from high throughput screening. In this talk, efforts, challenges and success in filtering the compounds to answer the long pending questions on lead-likeness and drug-likeness will be addressed. 
Biography 

Marcin J Skwark

University of Cambridge, UK

Title: CHOPIN, MABELLINI and HANSEN-Enabling combatting emerging antibiotic resistance in mycobacteria through multi-dimensional structural proteome data resources
Speaker
Biography:

Marcin J Skwark has completed his PhD in Biochemistry from Stockholm University. He is currently working as a Research Associate in the Department of Chemistry at University of Cambridge, UK.
 

Abstract:

Here we propose a set of computational resources to inform experiments and facilitate drug discovery against Mycobacterium spp. Mycobacteria are etiological agents of some of the most notorious hazards to public health-tuberculosis (M. tuberculosis) and leprosy (M. leprae). They are also responsible for opportunistic infections in Cystic Fibrosis (CF) patients (M. abscessus). Finally, they are conjectured to be the underlying cause of ulcerative colitis and Crohn’s disease (M. avium subsp. paratuberculosis). Despite their importance for public health, there are very few successful treatment strategies against Mycobacteria, due to their intrinsic resistance to antimicrobials, as well as inherent experimental difficulties in testing drug candidates in vitro. Our resources (CHOPIN: M. tuberculosis, MABELLINI: M. abscessus, HANSEN: M. leprae) provide comprehensive set of high-confidence structural models for the entire bacterial proteome. Models are generated using Vivace pipeline, using Fugue for template identification and Modeller for model building-both developed in-house. On contrary to typical protein structure prediction approaches (e.g. I-TASSER or HHpred), our approach aims to produce models that inform experiments and not necessarily maximize the stereo chemical quality and superposition to crystal structure. As our models are not over-optimized, they can be readily used for analysis of drug ability, effects of mutations and assessing interactions. By combining the results of state-of-art methods developed locally, with comprehensive survey of local and publicly available experimental results, CHOPIN, MABELLINI and HANSEN form resources of unprecedented utility-delivering results of rigorous computational analysis in a user-friendly, approachable and understandable way. The resources are free to use, constantly updated and produced in close collaboration with mycobacterial research community. All the software developed and used is open source and all the data is open access. 

Daniela C Vaz

University of Coimbra, Portugal

Title: Use of molecular interaction fields to understand drug resistance in HIV 1 protease caused by single point mutations
Speaker
Biography:

Daniela C Vaz has completed her Ph.D. in Biological Chemistry from the University of Coimbra. Her research focuses mainly on protein structure, folding, and stability, in relation to function and disease. She is currently working as a Professor at the School of Health Sciences of Leiria and is also a Member of the Coimbra Chemistry Centre at the University of Coimbra, Portugal.
 

Abstract:

Molecular Interaction Fields (MIF) is an archetypal computational chemistry technique that can be applied to capture a singular fingerprint of an ensemble of atoms on a protein and encode its physicochemical environment. Thus, MIFs have particular relevance in the context of binding hot spots and binding site analysis. Taking HIV 1 Protease (HIVPR) as case study, the present work focuses on a MIF-based in silico approach to achieve a qualitative interpretation and quantitative determination of mutation effects on HIVPR’s binding site, to help to understand translated changes in the enzyme’s structure and physicochemical environment. Assuming that binding sites with similar chemical environments have similar affinity for inhibitors, our method calculates and compares MIF similarities, visually assessing structural differences and quantifying their overlap through a Tanimoto coefficient. To assess the method’s ability to capture mutation induced chemical perturbations within HIVPR’s binding site, we collected 48 X-ray structures from the Protein Data Bank (PDB), from HIV strains either resistant or susceptible to protease inhibitors and quantified their binding site MIF similarities against a high quality, susceptible, reference structure. We observed and defined a threshold that discriminated most susceptible and resistant structures, confirming the MIF's suitability for our approach. Subsequently, we built homology models containing different reported single point resistance-conferring mutations using a single high-quality PDB structure as template. Root-Mean-Square Deviation (RMSD) values between template and model structures were calculated on residue by residue basis, confirming that the mutation was the only structural change. Then, the MIF similarities were determined, showing that this technique effectively captured subtle changes on HIVPR’s binding sites induced by the studied mutations. Along with the perspective of following an equivalent ligand based approach, we believe our results can be a promising starting point for developing an algorithm with drug resistance predictive power.
 

Daniela C Vaz

University of Coimbra

Title: RGA & GAI: On the characterization of two DELLA plant growth repressor proteins
Speaker
Biography:

Daniela C Vaz has completed her PhD in Biological Chemistry from the University of Coimbra. Her research focuses on protein structure, folding and stability in relation to function and disease. She is currently working as a Professor at the School of Health Sciences of Leiria and is also a Member of the Coimbra Chemistry Centre at the University of Coimbra, Portugal.
 

Abstract:

DELLA proteins are a family of nuclear proteins responsible for plant growth modulation. They act as growth repressor proteins in response to gibberellin signaling pathways. Five DELLA protein homologs were found in Arabidopsis thaliana, namely, RGA (Repressor of Gibberellic Acid), GAI (Gibberellic Acid Insensitive) and three RGA-like proteins (RGL-1, RGL-2 and RGL-3). The RGA- DELLA and GAI-DELLA protein homologs have been classified as Intrinsically Unstructured Proteins (IUPs) that undergo a disorder-to-order transition upon receptor binding. This structural change has found to be physiologically relevant for biological signaling and molecular recognition. Thus, in order to better characterize the structural features and molecular changes that govern these conformational variations of the polypeptide chain, we have produced recombinant RGADELLA and GAI-DELLA proteins in three length-versions, i.e. full-length, N-terminal and C-terminal versions. Full-length and terminal versions present different sequence motifs, attributed to different biological functions. All proteins were analyzed spectro-photometrically, via Light Scattering (LS), Circular Dichroism (CD) and intrinsic and extrinsic fluorescence (ANS binding), in order to compare spectral profiles, secondary structure propensities, levels of solvent exposure and structural compactness. Full-length and terminal variants exhibit different behaviors, spectral profiles and levels of compactness that can be related to different protein domains and ultimately to different functional implications.
 

Premkumar Dinadayala

Sanofi Pasteur

Title: Structural and functional characterization of various forms of glycated heat labile toxin protein
Biography:

Premkumar Dinadayala has completed his PhD from University of Toulouse and Postdoctoral studies from Colorado State University, Colorado. He is working as a Scientist since 12 years at Sanofi Pasteur. He has published more than 10 papers in journals.
 

Abstract:

Enterotoxigenic Escherichia coli (ETEC) are responsible for a high diarrheal disease burden, especially in children living in endemic countries and travelers visiting those countries. After oro-fecal transmission, ETEC reaches the small intestine where adhesion occurs through colonization factors. Then heat Labile Toxin (LT), one of the two enterotoxins produced by ETEC, is secreted and causes aqueous diarrhea. LT consists of five B sub-units, which are able to bind the monosialoganglioside GM1 and a single, catalytically-active A subunit stimulating the intracellular synthesis of cyclic Adenosine Monophosphate (cAMP), leading ultimately to fluid and electrolyte secretions into the intestinal lumen. Herein we characterized various purified forms of LT: (1) recombinant B subunit of LT (rLTB) (2) native LT purified from ETEC strain (nLT) and (3) recombinant LT purified from E. coli expressing the protein (rLT). SDS-PAGE analysis showed a difference of migration between the different LT forms confirmed by liquid chromatography coupled to MS (mass shift of 162 Da). This modification was found to be due to the glycation of LT subunits by galactose, a reducing sugar that is used in the LT purification process and remains present during the LT Lyophilization process. This observation has to be taken into account for the purification and storage of LT. Experiments are ongoing to determine if LT glycation could have an impact on the functional activity of LT using an in vitro assay based on cAMP release by epithelial cell line.