Understanding protein folding, evolution and function via molecular simulation

National Institute Of Diabetes And Digestive And Kidney Diseases

Investigators

Linked publications, trials & patents

Abstract

The project has addressed the following areas in the past year: 1. Complex coacervation of highly charged intrinsically disordered proteins. Recent work in collaboration with Ben Schuler's single molecule FRET group in Zurich has shown that high affinity disordered complexes of proteins or proteins and nucleic acids of opposite charge may be ubiquitous in cell nuclei. In addition, we have now found that some of these molecules undergo complex coacervation. Using novel multiscale simulation methodology, we are performing all-atom simulations of a complex coacervate of two nuclear proteins in order to elucidate the interactions responsible for stabilizing these phases, and we are also using new experimental data from FRET experiments and osmotic pressure measurements to help refine the force fields for salt bridges in the proteins as well as interactions of the proteins with ions (M. Ivanovic). (1) 2. Development of better coarse-grained models single-stranded nucleic acids for studying their phase separation/complex coacervation. We have successfully developed a coarse grained model for dsDNA for modeling its role in coacervation. We are now extending this to single stranded nucleic acids. For the purpose of developing better single-stranded nucleic acid models, we are using single-molecule FRET data and small-angle X-ray scattering data to do a force-balance optimization of the force-field. We are currently extending this model to include sequence-specific effects as well as back-mapping to atomistic simulations in a multiscale approach. (K. Lebold) 3. Development of transferable sequence-specific models for liquid-liquid phase separation (LLPS) of intrinsically disordered proteins. We had used a FRET data set on a set of proteins with diverse sequences collected under identical conditions in the Schuler lab to further refine our existing potential, resulting in significant improvements. Because coarse-grained models are necessarily a function of the experimental conditions, we are working to include such effects, in particular temperature as well as cosolvent or Hofmeister effects. (T. Dannenhoffer-Lafage) 4. Sequence dependence of co-phase separation of proteins. In cells, membraneless organelles do not simply consist of a single macromolecular component (or two, in the case of complex coacervation), but likely involve many different molecules. As a starting point for understanding the relative affinity one protein for a coacervate formed by another, we are collaborating with Tuomas Knowles at the University of Cambridge to develop a high-throughput sequence-scanning methodology, which we will use in conjunction with coarse-grained simulations to study this problem (2). (L. Good) 5. Inclusion of secondary structure in coarse-grained models. A common deficiency of many simple coarse grained models (such as single-bead models used for phase separation) as well as higher resolution coarse-grained models is the ability to form secondary structure based on sequence rather than as an input parameter of the simulation. We are developing sequence-specific model to capture secondary structure in coarse-grained models that will allow them to better describe transient secondary structure formation in disordered proteins, as well as formation of more extended secondary structure such as amyloids (R. Best). 6. Using sequence-based energy functions to describe protein fitness landscapes and for protein design. We have shown that it is possible to design novel foldable protein sequences using coevolutionary models. Most recently we have found that we can achieve increased thermostability via this route (2). We are also looking to develop similar ideas to identify proteins which naturally switch folds (such as RfaH), using sequence information. Separately, we are exploring the use of machine learning to predict fitness landscapes for proteins of any given structure; we are also exploring the ability of such models to capture the mechanism of allosteric transitions (L. Frechette, D. Wang). 7. Modelling properties of the extracellular matrix using coarse-grained models. We have recently started developing bottom up coarse-grained simulation models to describe the extracellular matrix and how they are related to its underlying molecular structure. We have initially focused on developing accurate atomistic models for collagen, and validating them against the available data from NMR and small angle X-ray scattering; we are extending this to coarse-grained models that will allow the investigation of packing of tropocollagen in collagen fibers. (G. Pantelopulos). 8. We are working in collaboration with Steve Vogel to understand the mechanisms in certain fluorescent proteins that allow coherent energy transfer via models parameterized from molecular simulations. (G. Taumoefolau). Group members or jointly supervised external collaborators involved in each project are listed at the end of each section.