BEGIN:VCALENDAR VERSION:2.0 PRODID:-//CERN//INDICO//EN BEGIN:VEVENT SUMMARY:Short Talk 11\, Raminta Venskutonytè - UrdA: structural character ization of a novel enzyme DTSTART;VALUE=DATE-TIME:20191011T080000Z DTEND;VALUE=DATE-TIME:20191011T082000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-772@lindico453.srv.lu.se DESCRIPTION:Speakers: Raminta Venskutonytė (Medical Structural Biology\, Lund University)\nUrocanate reductase (UrdA) is a bacterial enzyme that wa s first characterized in\n2012 and shown to reduce urocanic acid resulting in a product imidazole propionate\n(1). Unlike similar enzymes fumarate r eductases\, UrdA hasn’t been well investigated.\nBesides being an intere sting novel enzyme enabling bacteria to grow in anaerobic\nconditions with urocanic acid as electron acceptor (1)\, UrdA was shown to play a\nsignif icant role in human gut microbiota\, as imidazole propionate levels are\ni ncreased in people with type 2 diabetes and it further affects glucose met abolism\n(2).\nTwo domain construct of UrdA\, consisting of a FAD binding and a mobile domain\nwere successfully expressed\, purified and crystalliz ed. Four X-ray structures were\nobtained depicting different states of the enzyme: ADP bound\, FAD bound\,\nsubstrate/FAD bound and in complex with product/FAD. The data reveals the overall\nstructural arrangement of the e nzyme as well as the substrate binding mode and\nconformational changes.\n The role of UrdA in imidazole propionate production in relation to type 2 diabetes\nmakes the first structure of the UrdA of particular importance t o our understanding of\nthis enzyme.\n\nReferences\n1. Bogachev\, A. V. et al. (2012)\, Urocanate reductase of Shewanella. Molecular\nMicrobiology\, 86: 1452-1463.\n2. Koh A. et al. (2018) Microbially Produced Imidazole Pr opionate Impairs Insulin\nSignaling through mTORC1. Cell 175: 947-961.\n\n https://lindico453.srv.lu.se/event/125/contributions/772/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/772/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 9\, Annika Söderholm - Structure and substrate specifi city of the tryptophan biosynthesis enzyme IGPS from Pseudomonas aeruginos a DTSTART;VALUE=DATE-TIME:20191010T143000Z DTEND;VALUE=DATE-TIME:20191010T145000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-768@lindico453.srv.lu.se DESCRIPTION:Speakers: Annika Söderholm (Uppsala university)\nIn bacteria\ , tryptophan synthesis is performed by the enzymes encoded in the trp oper on. The product of the\ntrpC gene\, indole-3-glycerol phosphate synthase ( IGPS) catalyzes the indole-forming reaction of tryptophan\nsynthesis. The reaction mechanism includes a decarboxylation step of the substrate 1-(o-c arboxyphenylamino)\n1-deoxyribulose 5-phosphate (CdRP). The decarboxylatio n has been assumed to constitute an essential step\nof the mechanism since no activity with the decarboxylated variant of the substrate\, phenylamin odeoxyribulosephosphate\n(PAdRP)\, was observed in an early study on IGPS from Escherichia coli (Smith and Yanofsky\,\n1962).\nIn this study\, we de monstrate enzyme-catalyzed formation of the native product IGP from decarb oxylated\nsubstrate PAdRP using IGPS from Pseudomonas aeruginosa. Moreover \, the crystal structure of P. aeruginosa\nIGPS in complex with a substrat e analogue was solved to 2.1 Å resolution. By structural comparison to E. coli\nIGPS (Wilmanns et al.\, 1992)\, we provide structure-based hypothese s on the difference in substrate specificity\nbetween the E.coli and P. ae ruginosa homologs.\n\nReferences:\nSmith\, B. O. H. and Yanofsky\, C. (196 2) ‘Enzymes Involved in the Biosynthesis of Tryptophan’\, Methods Enzy mol.\,\n5\, pp. 794–806.\nWilmanns\, M. et al. (1992) ‘Three-dimension al structure of the bifunctional enzyme phosphoribosylanthranilate\nisomer ase: Indoleglycerolphosphate synthase from Escherichia coli refined at 2.0 Å resolution’\, Journal\nof Molecular Biology\, 223(2)\, pp. 477–507 .\n\nhttps://lindico453.srv.lu.se/event/125/contributions/768/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/768/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 7\, Ryan Oliver - Integrative structural biology with c omplementary experimental methods to describe viral capsid protein self-as sembly DTSTART;VALUE=DATE-TIME:20191010T101000Z DTEND;VALUE=DATE-TIME:20191010T103000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-765@lindico453.srv.lu.se DESCRIPTION:Speakers: Ryan Oliver (Dept of Biochemistry and Structural Bio logy)\nThe genetic code of viruses\, RNA or DNA\, are typically protected in an icosahedral capsid\, which is primarily\nassembled from over a hundr ed subunits of the same protein in a spontaneous self-assembly process. Si milar\nhighly efficient assembly processes are ubiquitous in biological sy stems\; viral capsids present a unique platform\nto exploit for therapeuti c advances in the targeted cellular delivery of cargo packaged within the capsid.\nOur research aims to provide a more detailed understanding of how this precise viral capsid protein assembly\nprocess occurs from a pool of single building blocks\, and additionally the effect and organization of nucleic\nacid present during assembly. Here\, we present results from smal l-angle neutron scattering experiments using\ncontrast variation to reveal the final assembled structural organization of both the protein and nucle ic\nacid components from recombinant Hepatitis B virus (HBV) capsid protei n and a synthetically prepared RNA\ncontaining the capsid protein binding domain. These data revealed that RNA was localized along the inner capsid\ nsurface. Time-resolved small-angle x-ray scattering (SAXS) experiments we re also used to determine the\nstructure during HBV capsid assembly in the presence and absence of RNA. We employed Bayesian statisticsbased\ncomput ational methods to extract kinetic parameters of assembly and the overall size and shape of the\ndominant structural intermediates from the SAXS dat a. Additional single-particle cryoEM reconstructions are\nprovided to asse ss the effect of RNA on the resulting assembled capsid structure. The comb ination of timeresolved\nscattering data\, Bayesian statistics\, and cryoE M structural analysis\, provides a framework which not\nonly describes the viral self-assembly process\, but can be extended to other hierarchical a ssemblies in biology.\n\nhttps://lindico453.srv.lu.se/event/125/contributi ons/765/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/765/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 6\, Mikael B. L. Winkler - STRUCTURAL INSIGHT INTO EUKA RYOTIC STEROL TRANSPORT THROUGH NIEMANN-PICK TYPE C PROTEINS DTSTART;VALUE=DATE-TIME:20191010T091000Z DTEND;VALUE=DATE-TIME:20191010T093000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-763@lindico453.srv.lu.se DESCRIPTION:Speakers: Mikael Winkler (Aarhus University)\nNiemann-Pick typ e C (NPC) proteins are essential for sterol homeostasis\, believed to driv e sterol integration into the vacuolar/lysosomal membrane before redistrib ution to other cellular membranes. Using a combination of crystallography\ , cryo-electronmicroscopy\, biochemical and in vivo studies on the Sacchar omyces cerevisiae NPC system\, NCR1/NPC2\, we recently generated a framewo rk for sterol membrane integration (Winkler et al.\, (2019)). Sterols are transferred between hydrophobic pockets of vacuolar NPC2 and membrane-prot ein NCR1. NCR1 has its N terminal domain (NTD) positioned to deliver a ste rol to a tunnel connecting NTD to the luminal membrane leaflet 50 Å away. A sterol is caught inside this tunnel during transport\, and a proton-rel ay network of charged residues in the transmembrane region is linked to th is tunnel supporting a proton-driven transport mechanism. We propose a mod el for sterol integration which clarifies the role of NPC proteins in this essential eukaryotic pathway and which rationalizes mutations in patients with Niemann-Pick disease Type C that I will present at the talk.\n\nWink ler et al\, Structural insight into eukaryotic sterol transport through Ni emann-Pick Type C proteins\, Cell (accepted\, 2019)\n\nhttps://lindico453. srv.lu.se/event/125/contributions/763/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/763/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 5\, Ronnie Berntsson - Structural insight into Gram-pos itive Type 4 Secretion Systems DTSTART;VALUE=DATE-TIME:20191010T074000Z DTEND;VALUE=DATE-TIME:20191010T080000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-761@lindico453.srv.lu.se DESCRIPTION:Speakers: Ronnie Berntsson (Umeå University)\nMultidrug resis tance in bacteria\, originating from conjugative gene transfer\, is an inc reasingly common problem\nin today’s world. The majority of bacteria tha t causes hospital infections are of gram-positive origin\,\nbut so far ver y little is known about their conjugation systems. To remedy this\, we aim to determine the\nmolecular structure and function of conjugation complex es belonging to Type IV Secretion Systems (T4SSs)\nfrom gram-positive bact eria. This will lead to a deeper insight into one of the main processes re sponsible for\nhorizontal gene transfer events\, including the spread of a ntibiotic resistance genes in bacteria.\nWe study the proteins involved in forming the T4SS biochemically\, structurally and biophysically. Since gr ampositive\nT4SSs are very dissimilar from their gram-negative counterpart s\, little can be deduced from the few\ngram-negative systems so far studi ed. Furthermore\, they occur in a number of pathogens\, such as enterococc i\,\nstreptococci and staphylococci. Another aspect that makes gram-positi ve T4SSs interesting is that they are\nused to efficiently transfer not on ly antibiotic resistance\, but also virulence factors.\nThese megadalton s ized systems are built up by i) extracellular adhesion proteins\, ii) memb rane channel proteins\nand iii) intracellular DNA processing proteins. Her e\, I will present our current understanding of the\nT4SS originating from the conjugative plasmid pCF10 of Enterococcus faecalis. Our work on this system\ncombines molecular biology\, biochemistry\, X-ray crystallography and Electron Microscopy. This has so far\nallowed us to determine structur es and understand some of the functions of the adhesion proteins as well a s\npart of the DNA processing proteins\, which will highlight both major d ifferences and similarities between the\ngram-positive and gram-negative s ystems.\n\nhttps://lindico453.srv.lu.se/event/125/contributions/761/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/761/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 2\, Sergei Grudinin - Novel algorithms for integrative structural biology. DTSTART;VALUE=DATE-TIME:20191009T121500Z DTEND;VALUE=DATE-TIME:20191009T123500Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-754@lindico453.srv.lu.se DESCRIPTION:Speakers: Sergei Grudinin (Inria / CNRS)\nIn my talk I will pr esent our approach for modeling macromolecular\nflexibility of large molec ular assemblies and how it can be combined with\nsparse experimental data obtained with small-angle and cross-linking\nexperiments.\nLarge macromole cular machines\, such as proteins and their complexes\, are\ntypically ver y flexible at physiological conditions\, and this flexibility is\nimportan t for their structure and function. Computationally\, it can be often\napp roximated with just a few collective coordinates\, which can be computed\n e.g. using the Normal Mode Analysis (NMA). NMA determines low-frequency\nm otions at a very low computational cost and these are particularly\nintere sting to the structural biology community because they are commonly\nassum ed to give insight into protein function and dynamics [1].\nOne of the cha llenges in the community is the explanation of solution smallangle\nscatte ring profiles. Very recently\, we designed a computational scheme\nthat us es the nonlinear normal modes [2] as a low-dimensional representation\nof the protein motion subspace and optimizes protein structures guided by the \nSAXS and SANS profiles [3\,4]. For example\, in the CASP12 and CASP13\ne xercises\, this scheme obtained best models for some (3 out of 9 in CASP12 )\nSAXS-assisted targets [5\,6]. Overall\, the flexible fitting scheme typ ically allows\na significant improvement of the goodness of fit to experim ental profiles in a\nvery reasonable computational time. The NMA analysis also allows to\nautomatically split macromolecules into rigid domains\, or to be used together\nwith the cross-linking data\, as we demonstrated in the recent CASP13\nchallenge [7].\n\nReferences:\n[1] Grudinin\, S.\, Lain e\, E.\, & Hoffmann\, A. (2019). Predicting protein functional\nmotions: a n old recipe with a new twist. bioRxiv\, 703652.\n[2] Hoffmann\, A. & Grud inin\, S. (2017). J. Chem. Theory Comput. 13\, 2123 –\n2134. For more in formation https://team.inria.fr/nano-d/software/nolbnormal-\nmodes/\n[3] G rudinin\, S. et al. (2017). Acta Cryst. D\, D73\, 449 – 464. For more\ni nformation https://team.inria.fr/nano-d/software/pepsi-saxs/\n[4] https:// team.inria.fr/nano-d/software/pepsi-sans/\n[5] http://predictioncenter.org /casp13/zscores_final_assisted.cgi?target_flag=S\n[6] Tamò\, G. E.\, Abri ata\, L. A.\, Fonti\, G.\, & Dal Peraro\, M. (2018). Proteins:\nStructure\ , Function\, and Bioinformatics\, 86\, 215-227.\n[7] http://predictioncent er.org/casp13/zscores_final_assisted.cgi?target_flag=X\n\nhttps://lindico4 53.srv.lu.se/event/125/contributions/754/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/754/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 3\, Prof. Richard Neutze: Time-resolved diffraction experi ments at X-ray free electron lasers reveal ultrafast structural changes in photosynthesis DTSTART;VALUE=DATE-TIME:20191009T144500Z DTEND;VALUE=DATE-TIME:20191009T152500Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-758@lindico453.srv.lu.se DESCRIPTION:Speakers: Richard Neutze (Gothenburg University)\nX-ray free e lectron lasers (XFEL) have sparked the development of time-resolved serial femtosecond crystallography (TR-SFX)\, which is a completely new experime ntal approach to understanding protein structural dynamics. We have used T R-SFX at the LCLS (an XFEL in California) to probe light-driven structural changes from picoseconds to microseconds in a bacterial photosynthetic re action centre. These integral membrane proteins harvest sunlight in order to transfer electrons from a special pair of bacteriochlorophylls to quino ne molecules that are located on the opposite side of an energy transducin g biological membrane. Coupled redox reactions balance the charges and thi s leads to a net effect of two pumped protons per photon absorbed. TR-SFX studies at the LCLS revealed structural changes on the picosecond time-sca le near the special pair (which is photo-oxidized by light) and the tightl y bound menaquinone (which accepts an electron from the special pair). The se structural results provide novel chemical insight into how protein stru ctural dynamics are able to help to stabilize the charge separated state. With the extension of serial crystallography to synchrotron radiation sour ces\, I argue that time-resolved diffraction studies will become more comm on in the future as new approaches allow new biological systems to be prob ed.\n\nhttps://lindico453.srv.lu.se/event/125/contributions/758/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/758/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 10\, Prof Erik Lindahl: Deciphering Allosteric Modulation in Ligand-Gated Ion Channels with Simulations\, X-ray crystallography\, Cr yo-EM and Neutron Scattering DTSTART;VALUE=DATE-TIME:20191011T091000Z DTEND;VALUE=DATE-TIME:20191011T095000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-774@lindico453.srv.lu.se DESCRIPTION:Speakers: Erik Lindahl (Stockholm University)\nLigand-gated io n channels control the electrical excitation of nerve cells\, in particula r in the post-synaptic membrane in response to chemical signals mediated b y neurotransmitters. These receptors exhibit an amazing diversity in detai led structure and function - some human channels have 15-20 slightly genes \, and with five subunits this can theoretically lead to almost a million different oligomers. They are further characterised by adopting both close d\, open and desensitised states - and in addition to the neurotransmitter s causing normal opening they are subject to secondary control - allosteri c modulation - by a number of drugs such as alcohols\, benzodiazepines\, n eurosteroids\, and anaesthetics that either potentiate or inhibit the agon ist response. I will present our work on understanding the molecular mecha nisms of these channels by using a broad range of experimental and theoret ical methods\, and illustrate that while each method has many shortcomings their combination increasingly enable us to capture different timescales\ , features\, interactions and not least dynamics of important membrane pro teins. For ligand-gated ion channels in particular\, this has enabled us t o explain several key mechanisms\, including identifying the separate pote ntiating and inhibitory binding sites\, showing how we can reverse the all osteric modulation of specific channels\, and propose detailed functional models even from intermediate-resolution structural data.\n\nhttps://lindi co453.srv.lu.se/event/125/contributions/774/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/774/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 6\, Dr. Andrey Gruzinov: Small-angle X-ray scattering (SAX S) for integrative structural biology DTSTART;VALUE=DATE-TIME:20191010T093000Z DTEND;VALUE=DATE-TIME:20191010T101000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-764@lindico453.srv.lu.se DESCRIPTION:Speakers: Andrey Gruzinov (EMBL c/o DESY\, Hamburg\, Germany)\ nSmall-angle X-ray scattering (SAXS) is a powerful method in the studies o f solutions of biological macromolecules and nanostructured systems [1] al lowing one to analyze the structure of native particles and complexes and to rapidly assess structural changes in response to variations in external conditions. Dedicated high brilliance synchrotron beamlines and novel dat a analysis methods [2] significantly enhanced resolution and reliability o f the structural models provided by SAXS. Very important is the ability of SAXS to quantitatively characterize complicated systems and mixtures in n ative environments and to see the biomolecules in action by rapidly observ ing responses to changing physical and chemical conditions (e.g. upon pH o r temperature changes\, ligand binding etc). \nGiven the limited informati on content in the scattering data\, robust data analysis and modelling met hods are of major importance for broad applications of solution SAXS in bi ology. To reduce the ambiguity of interpretation\, SAXS is often combined with other structural methods like crystallography\, NMR and electron micr oscopy\, and also with computational\, biophysical and biochemical techniq ues to build hybrid models. In classical applications\, SAXS generally yie lds low resolution quaternary structure but\, very importantly\, the metho d can also help to analyze equilibrium mixtures and to visualize flexible portions of the structures\, not seen by the high resolution methods. \nIn the present talk\, modern methods for SAXS data analysis will be presente d and illustrated by applications to characterize structures and conformat ional transitions of biological macromolecules in solution. Recent develop ments including\, in particular\, in-line chromatography approaches [3] wi ll be elucidated and perspectives of the synergistic use of SAXS for integ rative modeling utilizing complementary methods will be discussed.\n\nhttp s://lindico453.srv.lu.se/event/125/contributions/764/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/764/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 4\, Prof. Yvonne Jones: From structure to mechanism in the assembly and modulation of cell surface signalling complexes. DTSTART;VALUE=DATE-TIME:20191010T070000Z DTEND;VALUE=DATE-TIME:20191010T074000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-760@lindico453.srv.lu.se DESCRIPTION:Speakers: Yvonne Jones (Univ. of Oxford\, UK)\nIn my laborato ry we combine crystallographic\, biophysical\, electron and light microsco py based approaches into integrated structural biology analyses to study t he assembly and modulation of cell surface signalling complexes involved i n development and tissue homeostasis. We aim to generate mechanistic insig hts\, at atomic resolution\, which can be tested by functional studies in vitro and in vivo. I will discuss some of the recent results we have gener ated by applying this approach to the signalling mechanism of the semaphor in-plexin cell guidance system and to the extracellular modulation of sign alling by the morphogen Wnt. Published examples of our work on these two s ystems include the following:\n\nD. Rozbesky*\, R.A. Robinson\, V. Jain\, M. Renner\, T. Malinauskas\, K. Harlos\, C. Siebold\, and E.Y. Jones*. (20 19) Diversity of oligomerization in Drosophila semaphorins suggests a mech anism of functional fine-tuning. Nature Commun. 10\, 3691.\n\nY. Kong=\, B .J.C. Janssen=\, T. Malinauskas\, V.R. Vangoor\, C.H. Coles\, R. Kaufmann\ , T. Ni\, R.J.C. Gilbert\, S. Padilla-Parra\, R.J. Pasterkamp* and E.Y. Jo nes* (2016) ‘Structural basis for plexin activation and regulation.’ N euron 91\, 548-560\n\nS. Kakugawa=\, P.F. Langton=\, M. Zebisch=*\, S. How ell\, T.-H. Chang\, Y. Liu\, T. Feizi\, G. Bineva\, N. O'Reilly\, A.P. Sni jders\, E.Y. Jones* and J.-P. Vincent*. (2015) ‘Notum deacylates Wnts to suppress signalling activity.’ Nature 519\, 187-192\n\nhttps://lindico4 53.srv.lu.se/event/125/contributions/760/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/760/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 5\, Prof Liz Carpenter: Using Structural biology of human membrane proteins to understand the causes of genetic diseases DTSTART;VALUE=DATE-TIME:20191010T080000Z DTEND;VALUE=DATE-TIME:20191010T084000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-762@lindico453.srv.lu.se DESCRIPTION:Speakers: Liz Carpenter (University of Oxford\, UK)\nThe human genome project and subsequent sequencing efforts of thousands of patients and healthy individual provides a wealth of associations between variants in genes and diseases. At the SGC our aim is to create a step-change in d rug development by providing tools (proteins\, structures\, assays and bou nd small molecules)\, for proteins that are have associations with genetic disease. We focus in particular on proteins that are associated with neur opsychiatry\, cancer\, rare and metabolic disease\, as well as inflammator y conditions. These “Target Enabling Packages” or TEPs\, are made free ly available to advance our understanding of the biology of disease and to assist in the design of therapeutics. The Carpenter group focuses on inte gral membrane proteins\, including ion channels\, solute carriers\, ABC tr ansporters and enzymes. Here\, I will discuss three examples of genetic hi ts\, PKD2 in kidney disease\, TMEM16K in ataxia and DPAGT1 in congenital m yasthenia\, for which we have obtained structures\, and a wealth of additi onal improvement in our understanding disease biology. These examples of s tructures of genetic hits illustrate the power of structural biology\, as well as the need for extensive additional information\, to provide an unde rstanding of disease biology\, which is essential for development of thera peutics.\n\nhttps://lindico453.srv.lu.se/event/125/contributions/762/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/762/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 8\, Yong Wang - Integrative Ensemble Modeling of a Larg e Membrane Protein Complex Using Diverse and Ambiguous Information DTSTART;VALUE=DATE-TIME:20191010T141000Z DTEND;VALUE=DATE-TIME:20191010T143000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-790@lindico453.srv.lu.se DESCRIPTION:Speakers: Yong Wang (University of Copenhagen)\nMitochondria c ontain approximately 1200 different proteins\, 99% of which are synthesize d on cytosolic ribosomes and need to be delivered into the right destinati on through the intermembrane space by transport machineries\, such as the TIM chaperone. Currently\, the mechanistic and structural details of how t he TIM chaperone binds to these mitochondrial proteins remain elusive. To gain structural insight into the binding and chaperone mechanisms\, we foc used on the complex of the TIM9/10 chaperone and the mitochondrial GDP/GTP carrier membrane protein (Ggc1). Such complexes are difficult to study be cause they consist of a transiently formed\, dynamic complex between two f olded proteins and a membrane protein that should be solubilized and bound by the chaperone. X-ray crystallography has revealed the core structure o f the free chaperone protein\, but because of the dynamic nature and large size (~1400 amino acids) of the complex its structural features have rema ined elusive. Using an integrative approach that combines biochemical assa ys\, NMR spectroscopy and SAXS it was\, however\, able to obtain detailed but ambiguous information on the structures of the complex. In particular\ , the experiments showed that the complex consists of two well-structured (TIM9)3/(TIM10)3 hexamers bound to a mostly disordered Ggc1. In this work\ , we developed a protocol to integrate such heterogeneous experimental dat a with a coarse-grained molecular model to provide a description of the co nformational ensemble of the TIM9/10-Ggc1 complex. In particular\, we used a hybrid structure-based model (to describe the intra-molecular interacti ons within the folded chaperone)\, an NMR-derived contact potential for ch aperone-client interactions and a knowledge-based potential (to describe t he inter-molecular interactions between the chaperones and chaperone-clien t interactions). We used molecular dynamics (MD) simulations to sample the conformational landscape of the complex\, and the resulting coarse-graine d conformational ensemble was subsequently converted into all-atom resolut ion and refined using a Bayesian/Maximum Entropy re-weighting approach usi ng the SAXS data. This allows us to generate a weighted ensemble in agreem ent with experimental measurement. Such integrative structural modeling me thod is useful to generate a structural ensemble of large and dynamic prot eins in a both efficient and reliable way.\n\nReference:\nKatharina Weinh äupl\, Caroline Lindau\, Audrey Hessel\, Yong Wang\, Conny Schütze\, To bias Jores\,\nLaura Melchionda\, Birgit Schönfisch\, Hubert Kalbacher\, B eate Bersch\, Doron Rapaport\, Martha\nBrennich\, Kresten Lindorff-Larsen\ , Nils Wiedemann* and Paul Schanda*. Structural Basis of\nMembrane Protein Chaperoning Through the Mitochondrial Intermembrane Space. Cell\, 175\, 1 365-\n1379\, (2018)\n\nhttps://lindico453.srv.lu.se/event/125/contribution s/790/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/790/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 7\, Prof. Mei Hong: Structure and Dynamics of Amyloid Prot eins from Solid-State NMR: Glucagon & Tau DTSTART;VALUE=DATE-TIME:20191010T120000Z DTEND;VALUE=DATE-TIME:20191010T124000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-766@lindico453.srv.lu.se DESCRIPTION:Speakers: Mei Hong (MIT\, USA)\nProtein misfolding into amyloi d fibrils is common not only in neurodegenerative diseases but also in pha rmaceutical sciences\, where many peptide-based drugs have the tendency to fibrillize\, thus impeding solution formulation of the drug. Using solid- state NMR spectroscopy\, we have investigated the structure and dynamics o f two amyloid fibrils\, one formed by the peptide hormone glucagon\, which is used to treat diabetic hypoglycemia\, and the other formed by the micr otubule-binding protein tau\, which is found in many neurodegenerative dis eases. The glucagon fibril structure is unique among all amyloid proteins known to date: the -sheet is antiparallel rather than parallel hydrogen -bonded\, contains two coexisting molecular conformations in a single ultr astructural morphology\, and has an extraordinary -strand length of 10 nm. The 1.7 Å resolution structure reveals many stabilizing interactions for the fibril\, thus suggesting future strategies for designing glucagon analogs that resist fibril formation. Compared to glucagon\, the 40 kDa fu ll-length four-repeat tau protein forms a much more complex amyloid fibril \, with the majority of the protein being dynamically disordered. Using an extensive set of multidimensional correlation solid-state NMR techniques\ , we have determined the repeat domains that constitute the -sheet core \, and show that this core has a single molecular conformation. This monom orphic nature for an in-vitro tau fibril is fully consistent with the mono morphic nature of brain-derived tau fibrils known to date\, suggesting tha t in vitro fibrillized tau is a good model for studying in vivo tau fibril s. Further\, the segments outside the rigid core\, which appear as a “fu zzy coat” in electron micrographs\, are heterogeneously dynamic. The rep eats excluded from the rigid core exhibits partial mobility and -sheet character\, while the proline-rich domains undergo large-amplitude anisotr opic motions. These results suggest the structure and dynamics of tau in d iseases such as progressive supranuclear palsy\, and open the path for des igning tau inhibitors and imaging agents.\n\nhttps://lindico453.srv.lu.se/ event/125/contributions/766/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/766/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 8\, Prof. Henning Tidow: Structural studies of integral me mbrane proteins using stealth carrier nanodiscs DTSTART;VALUE=DATE-TIME:20191010T145000Z DTEND;VALUE=DATE-TIME:20191010T153000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-769@lindico453.srv.lu.se DESCRIPTION:Speakers: Henning Tidow (Hamburg University)\nStructural studi es of integral membrane proteins (IMPs) are challenging\, as many of them are inactive or insoluble in the absence of a lipid environment. We pionee red an approach making use of fractionally deuterium labelled ‘stealth c arrier’ nanodiscs that are effectively invisible to low-resolution neutr on diffraction and enable structural studies of IMPs in a lipidic native-l ike solution environment. We show the potential of the method in a joint s mall-angle neutron scattering (SANS) and X-ray scattering (SAXS) study of the ATP-binding cassette (ABC) transporter protein MsbA solubilized in the stealth nanodiscs. The data allow for a direct observation of the signal from the solubilized protein without contribution from the surrounding lip id nanodisc. Not only the overall shape but also differences between confo rmational states of MsbA can be reliably detected from the scattering data \, demonstrating the sensitivity of the approach and its general applicabi lity to structural studies of IMPs. In a follow-up project\, we could also apply this method to investigate the structural basis for the activation of an essential Ca2+-pump by its regulator calmodulin.\nThis methodology c an be applied to other classes of integral membrane proteins and paves the way for low-resolution structure determination of IMPs in solution using both ab initio and rigid body analysis approaches.\n\nhttps://lindico453.s rv.lu.se/event/125/contributions/769/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/769/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 1\, Prof. Poul Nissen: Structure and Dynamics of Membrane Transport Proteins DTSTART;VALUE=DATE-TIME:20191009T111500Z DTEND;VALUE=DATE-TIME:20191009T115500Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-752@lindico453.srv.lu.se DESCRIPTION:Speakers: Poul Nissen (Aarhus University)\nUsing membrane prot ein crystallography\, small-angle scattering techniques\, and cryo-EM\, an d also a range of biochemical and biophysical methods such as electrophysi ology\, single-molecule FRET\, and molecular dynamics simulations\, we hav e obtained deep insight into the functional cycle of primary active transp orters of the P-type ATPase family. These transport ATPases are fundamenta l to physiology\, and malfunctions are linked to diseases such as neurolog ical and cardiovascular disorders. \nThe transmembrane gradients for the k ey cations Na+\, K+\, and Ca2+ are generated by Na+\,K+-ATPase and Ca2+-AT Pases. In brain\, Na+\,K+-ATPase activity accounts for an estimated 40-70% of total ATP hydrolysis and potentiates e.g. Na+ and K+ channels for thei r activity in action potentials\, membrane potential\, and Na+ coupled tra nsport of e.g. glucose\, metabolite\, neurotransmitters\, Ca2+ efflux\, pH and Cl- control. Ca2+-ATPases maintain steep calcium gradients\, internal Ca2+ stores\, and cytoplasmic free calcium at accurate levels that define and potentiate calcium signalling pathways. \nLipid flippases\, also of t he P-type ATPase family (P4-ATPases) maintain asymmetric lipid distributio ns in biomembranes. Their activity potentiates membrane dynamics\, but the structure and function of lipid flippases remained enigmatic until recent ly. We determined the first structures of lipid flippases using cryo-EM an d revealed at the same time a detailed insight into lipid recognition and autoregulation.\nThe talk will cover methodological approaches supporting the functional and mechanistic insight we have gained.\n\nhttps://lindico4 53.srv.lu.se/event/125/contributions/752/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/752/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 2\, Prof. Yifan Cheng: Single particle cryo-EM of membrane proteins DTSTART;VALUE=DATE-TIME:20191009T123500Z DTEND;VALUE=DATE-TIME:20191009T131500Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-755@lindico453.srv.lu.se DESCRIPTION:Speakers: Yifan Cheng (UC San Francisco)\nWith the technologic al breakthroughs in the past few years\, single particle cryo-electron mic roscopy (cryo-EM) has enabled rapid progresses in structure determination of integral membrane proteins\, particularly ion channels. The pace of str ucture determination of integral membrane proteins by single particle cryo -EM is unprecedented in structural biology. With such a rapid progress\, i t is also very critical to interpretation of cryo-EM density maps carefull y to ensure that interpretation is data-driven. I will discuss some practi cal examples to demonstrate the significance of careful interpretations of single particle cryo-EM density maps. \n\nFurthermore\, as a prominent ex ample in structural biology of membrane proteins\, structural studies of t ransient receptor potential (TRP) channel superfamily demonstrated nicely how technological breakthroughs impacts scientific discoveries. As an exam ple of our recent studies of TRP channels by single particle cryo-EM\, TRP V5 (transient receptor potential vanilloid 5) represents a unique calcium- selective TRP channel essential for calcium homeostasis. Unlike other TRPV channels\, TRPV5 and its close homolog\, TRPV6\, do not exhibit thermosen sitivity or ligand-dependent activation but are constitutively open at phy siological membrane potentials and modulated by calmodulin (CaM) in a calc ium-dependent manner. Structural studies of truncated and full-length TRPV 5 in lipid nanodiscs\, as well as of a TRPV5 W583A mutant and TRPV5 in com plex with CaM provide novel insights to the mechanism of calcium regulatio n and reveal a flexible stoichiometry of CaM binding to TRPV5.\n\nhttps:// lindico453.srv.lu.se/event/125/contributions/755/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/755/ END:VEVENT BEGIN:VEVENT SUMMARY:Keynote 9\, Prof Nieng Yan: How is electrical signal generated? Structural and mechanistic investigations of Nav channels DTSTART;VALUE=DATE-TIME:20191011T070000Z DTEND;VALUE=DATE-TIME:20191011T074000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-770@lindico453.srv.lu.se DESCRIPTION:Speakers: Nieng Yan (Princeton University\, USA)\nThe voltage- gated sodium (Nav) channels are responsible for the initiation and propaga tion of action potentials. Being associated with a variety of channelopath ies\, they are targeted by multiple pharmaceutical drugs and natural toxin s. We determined the crystal structure of a bacterial Nav channel NavRh in a potentially inactivated state a few years ago\, which is a homotetramer in primary sequence but exhibits structural asymmetry. Employing the mode rn methods of cryo-EM\, we determined the near atomic resolution structure s of a Nav channel from American cockroach (designated NavPaS) and from el ectric eel (designated EeNav1.4). Most recently\, we have determined the c ryo-EM structures of the human Nav channels\, Nav1.2\, Nav1.4\, and Nav1.7 in complex with distinct auxiliary subunits and toxins.These structures r eveal the folding principle and structural details of the single-chain euk aryotic Nav channels that are distinct from homotetrameric voltage-gated i on channels. Unexpectedly\, the two structures were captured in drasticall y different states. Whereas the structure of NavPaS has a closed pore and the four VSDs in distinct conformations\, that of EeNav1.4 and the human c hannels is open at the intracelluar gate with VSDs exhibiting similar “u p”states. The most striking conformational differenc occurs to the III-I V linker\, which is essential for fast inactivation. Based on the structur al features\, we suggest an allosteric blocking mechanism for fast inactiv ation of Nav channels by the IFM motif. Structural comparison of the confo rmationally distinct Nav channels provides important insights into the ele ctromechanical coupling mechanism of Nav channels and offers the 3D templa te to map hundredes of disease mutations.\n\nhttps://lindico453.srv.lu.se/ event/125/contributions/770/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/770/ END:VEVENT BEGIN:VEVENT SUMMARY:Concluding remarks DTSTART;VALUE=DATE-TIME:20191011T095000Z DTEND;VALUE=DATE-TIME:20191011T101000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-775@lindico453.srv.lu.se DESCRIPTION:Jens Lagerstedt\, Susanna Törnroth-Horsefield\, Thomas Ursby\ , Trevor Forsyth\, Christine Ziegler\n\nhttps://lindico453.srv.lu.se/event /125/contributions/775/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/775/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 12\, Magnus Andersson - Tracking Ca2+ ATPase Intermedia tes in Real-Time by X-ray Solution Scattering DTSTART;VALUE=DATE-TIME:20191011T085000Z DTEND;VALUE=DATE-TIME:20191011T091000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-773@lindico453.srv.lu.se DESCRIPTION:Speakers: Magnus Andersson (Umeå University)\nSarco/endoplasm ic reticulum Ca2+ ATPase (SERCA) transporters regulate calcium signaling b y active calcium\nion reuptake to internal stores. Several of the structur al transitions associated with transport have been characterized\nby X-ray crystallography\, but critical intermediates of the inward-outward switch ing are missing.\nWe combined time-resolved X-ray solution scattering (TR- XSS) experiments and molecular dynamics (MD)\nsimulations for real-time tr acking of concerted SERCA reaction-cycle dynamics in the native membrane. The\nTR-XSS pre-pulse model differed in the domain arrangement compared to Ca2E1 crystal structures. A 1.5\nms intermediate showed closure of the cy tosolic domains typical of Ca2+- and ATP-bound E1 states. A subsequent\ntr ansi-ent state with a 13 ms rise-time showed a novel actuator (A) domain a rrangement that exposes the\nADP-binding site after phosphorylation. Hence \, the obtained TR-XSS models determine the relative timing\nof so-far elu sive domain rearrangements in a native environment.\n\nhttps://lindico453. srv.lu.se/event/125/contributions/773/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/773/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 10\, Hongyi Xu - Solving the First Novel Protein Struct ure by Micro-Crystal Electron Diffraction DTSTART;VALUE=DATE-TIME:20191011T074000Z DTEND;VALUE=DATE-TIME:20191011T080000Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-771@lindico453.srv.lu.se DESCRIPTION:Speakers: Hongyi Xu (Stockholm University)\nMicro-crystal elec tron diffraction (MicroED) has shown in recent years to be a promising met hod for determining\nmacromolecular structures (1–5). It enables structu ral biologists to study proteins from micron-sized\n3D crystals that are t oo small to be studied by conventional X-ray crystallography. Furthermore\ , MicroED\ncan be applied to biomolecules of low molecular weight that are beyond what can so far be resolved by single\nparticle cryo-EM (6\,7). Ho wever\, up to now\, all protein structures determined by MicroED had alrea dy been\nsolved previously by X-ray crystallography. Here\, we present for the first time an unknown protein structure\n– an R2lox metalloenzyme – solved using MicroED (8). MicroED data were collected from plate-like crystals\nwith an average size of 2 μm × 2 μm × 0.5 μm. By overcoming challenges in sample handling\, cryo-EM specimen\npreparation\, limited d ata completeness and low signal-to-noise ratio\, we are able to solve the structure\nby molecular replacement with a search model of less than 36% s equence identity. The resulting electrostatic\nscattering potential map at 3.0 Å resolution is of sufficient quality to allow accurate model buildi ng and refinement\,\nproviding biologically relevant information on the en zyme. Our results demonstrate MicroED can be\nused for solving novel prote in structures\, using only standard X-ray crystallography software. These findings\nillustrate that electron crystallography has the potential to be come a widely applicable tool for revealing new\ninsights into protein str ucture and function\, opening up new opportunities for structural biologis ts.\n\nRefrences\n1. Shi\, D.\, Nannenga\, B. L.\, Iadanza\, M. G. & Gonen \, T. eLife 2\, (2013).\n2. Nannenga\, B. L.\, Shi\, D.\, Leslie\, A. G. W . & Gonen\, T. Nat. Methods 11\, 927–930 (2014).\n3. Yonekura\, K.\, Kat o\, K.\, Ogasawara\, M.\, Tomita\, M. & Toyoshima\, C. Proc. Natl. Acad. S ci. 112\, 3368–3373\n(2015).\n4. Clabbers\, M. T. B. et al. Acta Crystal logr. Sect. Struct. Biol. 73\, 738–748 (2017).\n5. Xu\, H. et al. Struct ure\, 26\, 667-675 (2018).\n6. Khoshouei\, M.\, Radjainia\, M.\, Baumeiste r\, W. & Danev\, R. Nat. Commun. 8\, 16099 (2017).\n7. Henderson\, R. Q. R ev. Biophys. 28\, 171 (1995).\n8. Xu\, H. et al. Sci. Adv. 5\, eaax4621 (2 019).\n\nhttps://lindico453.srv.lu.se/event/125/contributions/771/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/771/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 3\, Viktoria Bågenholm - Two GH26 β-mannanases from B acteroides ovatus: structure and role in galactomannan degradation DTSTART;VALUE=DATE-TIME:20191009T142500Z DTEND;VALUE=DATE-TIME:20191009T144500Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-757@lindico453.srv.lu.se DESCRIPTION:Speakers: Viktoria Bågenholm (Lund University)\nGalactomannan s are hemicelluloses composed of a β-1\,4-linked mannose backbone with α -1\,6-galactose substitutions.\nThey are part of our diet as seed storage polysacchardies and food thickeners and are utilised\nby several human gut bacteria (1). One such bacteria\, Bacteroides ovatus\, contains a gene cl uster encoding\ntwo glycoside hydrolase family 26 β-mannanases\, BoMan26A and BoMan26B (2). BoMan26B generates a\nrange of product lengths upon man nan hydrolysis\, prefers longer substrates and is less restricted by galac tose\nside-groups than BoMan26A\, which mainly generates mannobiose (3\,4) . The results suggest that BoMan26B\nperforms the initial attack on galact omannan\, generating oligosaccharides that are further hydrolysed by Bo-\n Man26A. Crystal structures of these two enzymes reveal the structural basi s for their biochemical differences.\nBoMan26B\, with galactosyl-mannotetr aose bound in subsites –5 to –2\, has an open and long active site cle ft\nwith W112 in subsite –5 concluded to be involved in mannosyl interac tion (4). Moreover\, K149 in the –4 subsite\ninteracted with the galacto syl side-group of the ligand\, which may indicate a preference in for subs tituted\nmanno-oligosaccharides (4). BoMan26A instead revealed a narrow ac tive site cleft that is restricted in one end\nby a loop\, explaining its preference for generating shorter products (6).\n\n1. Kulcinskaja\, E.\, A . Rosengren\, R. Ibrahim\, K. Kolenova and H. Stalbrand Appl Environ Micro biol (2013). 79:\n133-140\n2. Martens\, E. C.\, E. C. Lowe\, H. Chiang\, N . A. Pudlo\, M. Wu\, N. P. McNulty\, D. W. Abbott\, B. Henrissat\, H. J.\n Gilbert\, D. N. Bolam and J. I. Gordon PLoS Biol (2011). 9: e1001221\n3. B ågenholm\, V.\, S. K. Reddy\, H. Bouraoui\, J. Morrill\, E. Kulcinskaja\, C. M.\, Bahr\, O. Aurelius\, T. Rogers\,\nY.Xiao\, D. T. Logan\, E. C. Ma rtens\, N. M. Koropatkin and H. Stålbrand J Biol Chem (2017). 292: 229-24 3\n4. Bågenholm\, V.\, M. Wiemann\, S. K. Reddy\, A. Bhattacharya\, A. Ro sengren\, D. T. Logan and H. Stålbrand J\nBiol Chem (2019). 294: 9100-911 7\n\nhttps://lindico453.srv.lu.se/event/125/contributions/757/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/757/ END:VEVENT BEGIN:VEVENT SUMMARY:Working Group presentations 4X10 min DTSTART;VALUE=DATE-TIME:20191009T134500Z DTEND;VALUE=DATE-TIME:20191009T142500Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-756@lindico453.srv.lu.se DESCRIPTION:https://lindico453.srv.lu.se/event/125/contributions/756/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/756/ END:VEVENT BEGIN:VEVENT SUMMARY:Short Talk 1\, Veronika Nesverova - Structural insights into AQP2 targeting to multivesicular bodies DTSTART;VALUE=DATE-TIME:20191009T115500Z DTEND;VALUE=DATE-TIME:20191009T121500Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-753@lindico453.srv.lu.se DESCRIPTION:Speakers: Veronika Nesverova (Lund University)\nThe multivesic ular sorting machinery is a crucial mechanism for targeting membrane prote ins\nfor recycling or degradation. The lysosomal trafficking regulator-int eracting protein 5 (LIP5)\nwhich coordinates the action of this machinery is also known to bind directly to the membrane\nprotein cargo. In case of aquaporin 2 (AQP2) the binding of LIP5 during the endocytic pathway\nin ki dney collecting duct cells ensures an effective regulation of urine volume [1].\nIn our group\, we have previously studied the role of AQP2 phosphor ylation in AQP2-LIP5\ninteraction [2]. Currently we are focusing on elucid ating the structural details of the complex\nin order to better understand how membrane proteins are delivered to the multivesicular\nbodies. We hav e constructed alanine mutants of single residues in the proposed binding s ites\nof both AQP2 and LIP5. Studying the binding affinity of these mutant s using fluorescence\nquenching helps us understand which residues are dir ectly involved in the binding.\nFurther\, AQP2 was successfully incorporat ed into MSP-based nanodiscs and negative stain\nelectron microscopy confir med homogeneous state of the particles. We have collected high\nresolution images on Titan Krios and are currently processing the data.\n\n[1] B. W. M. Van Balkom\, M. Boone\, G. Hendriks\, E. Kamsteeg\, J. H. Robben\, H. C. Stronks\, A.\nVan Der Voorde\, and F. Van Herp\, “LIP5 Interacts with Aquaporin 2 and Facilitates Its\nLysosomal Degradation\,” pp. 990–100 1\, 2009.\n[2] J. V. Roche\, S. Survery\, S. Kreida\, V. Nesverova\, H. Am pah-Korsah\, M. Gourdon\, P. M. T.\nDeen\, and S. Törnroth-Horsefield\, “Phosphorylation of human aquaporin 2 (AQP2)\nallosterically controls it s interaction with the lysosomal trafficking protein LIP5\,” J. Biol.\nC hem.\, vol. 292\, no. 35\, pp. 14636–14648\, 2017.\n\nhttps://lindico453 .srv.lu.se/event/125/contributions/753/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/753/ END:VEVENT BEGIN:VEVENT SUMMARY:Introduction\, LINXS and Core Group of ISB DTSTART;VALUE=DATE-TIME:20191009T110000Z DTEND;VALUE=DATE-TIME:20191009T111500Z DTSTAMP;VALUE=DATE-TIME:20260524T211302Z UID:indico-contribution-125-751@lindico453.srv.lu.se DESCRIPTION:https://lindico453.srv.lu.se/event/125/contributions/751/ LOCATION:Kulturen Auditorium URL:https://lindico453.srv.lu.se/event/125/contributions/751/ END:VEVENT END:VCALENDAR