Thematic Meeting, Berlin 2017: Session I & Session II

Just a blink and this fantastic conference was over. Equally my time for writing some posts passed by, which preparation I fairly underestimated. So, now I would like to share my notes with you. Hope you enjoy reading my post-conference posts, to remember the great talks or get an impression, what you missed! Let’s start with session I & II!

After planning this meeting thoroughly together with Helen Berman, Andrea Cavalli and Gerhard Hummer for several years, Kresten Lindorff-Larsen addressed the opening remarks emphasizing what they aimed for this program and how much they looked forward to tackle it. And they were not alone! Finally, the first session on “Disordered Protein Ensembles” started, including talks by Adriaan Bax, Tanja Mittag, Teresa Head-Gordon and Paul Robustelli on Friday and Martin Blackledge and Birthe Kragelund on Saturday morning.
For me it was the first time I heard such comprehensive collection of talks about (intrinsically) disordered proteins (IDP) and this session had a great mixture of experimental methods to computational strategies and the benefit of their conjunction.

Starting with Adriaan Bax, he observed protein folding and misfolding by pressure jump NMR with special focus on HIV-1 Protease. The challenge in there is indeed the pressure but by managing this, he observed pressure-induced fully reversible unfolding including slow exchange time scale. Additionally, monomer exchange faster than stable dimer. He further looked into single residue cis-trans isomerisation. By investigating hydrogen bonding network stability, he sees a significant shift towards low pressure. For Abeta(1-40) fibril formation, real time NMR signals disappear after dropping pressure from 2.4 kbar to 1 bar, which is then heterogeneous, but rapidly reappears after jumping to high pressure. For his third system (Ubiquitin), the showed the influence of pressure by revealing an intermediate state, leaving open that this might also be due to mutations.

Next Tanja Mittag studied the effect of multi-site phosphorylation on conformations of intrinsically disordered proteins, exemplary S. cerevisiae transcription factor Ash1. Ash1 is expected to be collapsed and to expand upon multi-site phosphorylation but SAXS shows only little differences. By following an ensemble optimization method, she identified preferences for expanded conformations to be insensitive to screening of long-range electrostatic interactions, but reacting to the presence of weak local structural preferences. She identifies sequence features, mainly a relationship between proline and charged amino acids, deriving intrinsic sequence code expansion. She argues against chain compaction upon multi-site phosphorylation due to proline isomerization or presence of pSer/Arg or pThr/Arg salt bridges. Additionally, all-atom simulations could reproduce the experimental observations. Finally, she states that a large enough concentration of Prolines distributed along a sequence can buffer changes in net charge per residue while changes in local conformation occur.

After a refreshing coffee break, Teresa Head-Gordon presented new methods for generating and evaluating conformational ensembles. She looked at secondary structure propensities for Abeta42 and Abeta43 and observed differences in N-terminus and central hydrophobic core with REMD simulation, indicating either bad force-fields, poor sampling or both. There she introduced Temperature Cool-Waking (TCW) using annealed importance sampling and could indicate poor sampling. She states that TCW and polarizable forcefields work for folded proteins and are robust for IDPs, whereas Boltzmann priors may be questionable for IDPs. Monte-Carlo side chain ensemble (MC-SCE) has implemented a sophisticated energy function to distinguish different side chain packings based on Boltzmann factor. She concluded that MC-SCE and entropy expansions are informative about improved catalysis.

For the last talk of the first day, Raul Robustelli from D.E. Shaw Research gave insights in ongoing force field development, focusing on ordered and disordered protein states. The problem of current force fields for IDPs is that they tend to being structurally too compact relative to experiments. In this context, he stated the importance of an improved water model, as water dispersion energies have been systematically underestimated. Therefore, he introduced the TIP4P-D water model. Enhanced hydrogen bond potential and ‘fractional’ charges showed improved fits of non-bonded parameters to quantum data. As those analysis fit nicely, still further improvements have to be conducted, e.g. to challenge the over-representation of beta sheets.

With Paul Robustelli, an interesting first day at the “Conformational Ensembles from Experimental Data and Computer Simulations” conference ended and everybody went over to the welcome reception to enjoy a beautiful evening on the terrace outside. Perfect time for some networking. You can find some impression on twitter (@BiophysicalSoc).

Day two started with Martin Blackledges talk about “Large-scale Protein Conformational Dynamics from NMR and Molecular Simulation. From Fundamental Biophysics to Biological Function”. He aims to understand large-scale domain motion in influenza, where temperature is a key factor. Problematically is that its crystal conformation cannot bind to importin alpha. For NMR, its 2 domains do not interact with each other, rather having an open and a closed form. He finds that highly conserved salt bridges are involved in stabilizing the closed formation. He uses chemical exchange saturation transfer (CEST) to measure interconversion rate and population of substates. Using integrated structurally dynamics (NMR, smFRET, SAXS) CEST analyses are supported and dynamic interconversion between open and closed form of H5Na influenza 627 are revealed .
To explain how IDPs interact with their physiological partners, Martin introduced Asteroids, a selection tool of ensemble descriptions of intrinsically disordered systems, which was tested using target ensembles. In order to understand the mechanism of highly dynamic IDPs interactions, first their intrinsic dynamics have to be understood and therefore he developed a physical framework by comparing conformational sampling between 274 -298 K.
Using the ABSURD (average block selection using relaxation data) procedure, he could reproduce the experimental data, overcoming current limitations of MD simulations of IDPs. It identifies ensembles of trajectories of IDPs and can thereby map extent of inter-residue dynamic correlation.

The last talk for this session was held by Birthe Kragelund on dynamics and disorder in class 1 Cytokine receptors. She states that disorder is an important and integral part of membrane proteins as intrinsically disordered regions (IDRs) co-structure them as regulatory platforms with the need of structural information on bound state. Still, they are often not recognized. For membrane proteins, other players come into account. Disordered lipid binding motifs still need to be identified and understood, especially short linear motifs. In general, she combined experimental and computational techniques, including NMR, X-ray scattering, mass spectrometry, simulation and molecular modeling, solving the prolactin receptor monomeric structure. As the transmembrane domain is a weak dimer with highly specific interactions, its study is more challenging. She could identify two conformations, providing more insights into its dynamical function, e.g. that specificity can be modified by one single methyl group. Finally, she describes the multiple transmembrane domain states which are influenced by lipid composition, kinase binding and dimerization.

After the coffee break, the second session started on “Integrative and Hybrid Methods” with Andrej Sali, Alexandre Bonvin and Ji-Joon Song started.

Andrej Sali introduced us his research on integrative structure determination, where he uses experiments, physical theory and statistical inference to maximize accuracy, resolutions, completeness and efficiency. He presented 4 general steps of his integrative modeling platform (IMP), first naming gathering of information by experimental data, statistical inference and physical principles, second designing system representation and scoring, third followed by sampling and finally analysis and validation. Following this workflow for the Spindle Pole Body, he obtained a validated structure including insights into functional implications of the model.

Next, Alexandre Bonvin presented his integrative modeling platform HADDock (high ambiguity driven docking). It incorporates ambiguous and low-resolution data to aid the docking of up to 6 molecules and has a powerful algorithm to handle flexibility at the interface including refinement in explicit solvent. Using RNA-polymerase II, he introduced DisVis for explorative modeling and consistency quantification of information content of distance restraints solely based on geometric considerations. It can provide information about possible interfaces and calculable information to guide modeling but does not account for conformational changes and energetics.

Ji-Joon Song focused with his talk on the human importin4_histone H3/H4 Asfla complex, which is a perfect example for a successful integrative structural approach, as he was able to solve the whole complex by combining several techniques, such as x-ray crystallography, SAXS, EM, Mass spectrometry, biological application and modeling. He also stated that C-importin4 directly interacts with histon H3 peptide via a highly acid patch. The validation of the complex via single particle electron microscopy, small angle x-ray scattering and cross-linking mass spectrometry confirmed conformational flexibility.

Closing his talk, many discussions were extended during a manifold but time-wise rather tight lunch outside at the venue. Enjoying the sun we were waiting for the next session, curious what surprises there might be… You want to know what happened? Check out my next post on the third session about “Interpreting Experiments through Molecular Simulations”!

Cheers,

Johanna

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