[Colloquium] Fwd: Beagle - May 2018 Newsletter

Thu May 24 15:15:04 CDT 2018

>> Is this email displaying correctly? Try the web version <https://www.docdroid.net/EyMWW7I/may-newsletter-2018-finalodg-0.pdf>	Volume 20, May 2018
>> 
>> Science Highlight
>> Combining Computation and Experiment to Study Src Family Kinase Membrane Association
>> Matthew P Pond and Benoît Roux 
>> 
>> ABSTRACT 
>> Kinases are among the most crucial signaling proteins in the human body and hold key roles in essential cellular processes. The kinase superfamily is among the most well studied in nature, but kinases of the Src family have garnered special attention because of the well-documented relation between their aberrant signaling and cancer. Members of the family share a common architecture, comprising several Src-homology (SH) domains. Those include the SH3, SH2, and SH1 (the kinase proper) domains, which are linked to the membrane-anchoring SH4 domain through an intrinsically disordered region of low sequence conservation called the “Unique” (U) domain. Despite the wealth of information on this important family of proteins and the several known roles of the SH4-U in kinase regulation, we still do not know how this region interacts with the membrane or communicates with the rest of the protein. We are performing experimentally guided all atom molecular dynamics simulations on Beagle2 to investigate SH4-U–membrane interactions to provide a fundamental understanding of the SH4-U. We anticipate that these and other similar studies will lead to the discovery of new intermediate states and alternate inhibitory strategies of kinase activities for drug development.
>> 
>> INTRODUCTION 
>> Kinases are enzymes that regulate many key cellular processes via their roles in signal transduction pathways. c-Src kinase, the prototypical member of the nine Src family kinases (SFKs: Src, Yes, Fyn, Fgr, Lyn, Hck, Lck, Blk, and Yrk), is a non-receptor tyrosine kinase vital for cell growth, proliferation, metabolism, differentiation, adhesion, and migration. Over-activation of Src is often at the origin of tumorigenesis, tumor metastasis, and obesity. Anti-cancer drugs such as the kinase inhibitors dasatinib and bosutinib were designed to target Src specifically because of its importance in cellular signaling.
>> 
>> Hematopoietic cell kinase (Hck) is a phagocyte specific proto-oncogene of the Src family that plays a critical role in Bcr/Abl-chronic myeloid leukemia (CML); constitutive activation of Hck by direct interaction with the oncogene Bcr/Abl is required for the establishment of leucocyte transformation. More generally, Hck may play a role in promoting metastatic spread through its activation in the solid tumor micro-environment, and elevated expression levels of Hck have been observed in many cancers, with melanoma, ovarian, breast, and uterine being most prominent. Combined with the mild phenotype displayed by mice lacking Hck, the connection between Hck and cancer has led to a pharmacological interest in Hck as a rational target for cancer therapy, most notably in the fight against SFK-dependent resistance to imatinib in CML patients.
>> 
>> 
>> Figure 1. Cartoon representation of the p61 isoform of the Src Family Kinase Hck showing how the SH4 & Unique domains link the core of the protein to the membrane. (Bottom) Schematic diagram showing the relative sizes and locations of the different Hck domains.
>> Structurally, the members of the Src family of kinases share a common architecture, comprising several Src-homology (SH) domains. Those are the SH3, SH2, and SH1 (the kinase) domains, preceded by a ~80 residue region of low conservation called the Unique (U) domain and a membrane-targeting SH4 region at the N terminus (Figure 1). The topological organization of the full-length kinase is SH4-U-SH3-SH2-SH1. Crystal structures have shown that the catalytic activity of SFKs is tightly regulated by autoinhibition, with activation being achieved by displacing one or all of the interactions between the SH1 and the regulatory SH3 and SH2 domains. Acylation of the SH4 domain (myristoylation and palmitoylation) is critical for membrane association of SFKs, but little is known about the contributions of residues in the U domain to this process. In addition, clear lines of evidence establish that SH4-U domains influence SFK substrate specificity and function. Interestingly, the sequence of the Hck U domain is well conserved across different species, despite differences between SFK members, arguing for its importance as a critical structural feature for regulating SFK function. In fact, localization of activated SFKs at the membrane through the SH4-U region is critical for the regulation of specific cellular processes, and may permit a tight selection over downstream substrates. An atomic level characterization of SH4-U–membrane interactions is therefore of high importance and may to lead to the discovery of new intermediate states and alternate inhibitory strategies of kinase activities for drug development.
>> 
>> Exploring protein structure at the membrane interface is, however, a difficult task. Many experimental techniques used in modern biophysical studies require the molecule to be tumbling freely in solution (i.e., not attached to a membrane) or lack the resolution to provide information on the atomic scale. Furthermore, sequence analysis indicates that the U domain is intrinsically disordered; thus, one cannot simply expect the U domain to adopt a unique molecular structure. Molecular modeling is therefore necessary to provide missing information, but these techniques also face challenges for this class of proteins. Molecular dynamics (MD) is the computational method of choice for studying molecules of this scale, but MD force fields have generally been developed with the intention of exploring folded proteins with well defined tertiary structures, and consequently, these simulations are often unable to capture essential properties of disordered proteins. MD simulations also often rely on starting from crystal or NMR structures that represent energetically favorable states, thereby subverting the need to extensively sample to find relevant states of the protein. Without the benefit of these initial structures, it is possible that we may never locate important interactions between the protein and membrane with MD alone.
>> 
>> RESULTS 
>> To overcome these challenges, we have designed new tools to incorporate experimental information into MD simulations. Our recent work involves using Neutron Reflectometry (NR) data collected by the Lösche lab at Carnegie Mellon University/NIST. NR is a powerful experimental technique for investigating molecules at interfaces. When studying protein–membrane interactions, the experiments are conducted using lipid bilayers tethered to a fixed surface, which are placed in a flow cell chamber to allow for the controlled introduction of solution containing protein to the system. The technique only yields a low-resolution one dimensional density profile of how the different components in the system are organized, but it provides extremely helpful constraints for the MD simulations. To improve conditions and limit the complexity of the system, we chose to remove the SH4-U domains from the rest of the protein (HckSH4-U) to study it in isolation. We then transform the density profiles provided by NR into restraints for our MD simulations. Since HckSH4-U is expected to remain largely disordered upon membrane association, it is unreasonable to assume that one copy of the protein will be able to fit the density profile obtained by NR; therefore, we use multiple copies of the system in which the restraints are applied to the ensemble instead of a single copy. The cost comes at increasing the system size, and even with the modest size of HckSH4-U, after explicit water and membrane molecules are added to the multiple copies, we have nearly 6.5 million atoms in total. The results from the simulations run on Beagle2 can be seen in Figure 2, and give the first glimpse into HckSH4-U–membrane association at the atomic scale. The structures identify previously unknown Hck-membrane interactions and provide testable hypothesis for further experimentation.
>> 
>> 
>> Figure 2. Results from the MD simulations showing the ensemble of conformations representing the experimental data. For presentation purposes, the lipids are represented as spheres located at the location of phosphorus atoms, and water molecules have been removed.
>> Additional information about science on Beagle can be found here: Beagle2 Website <http://beagle.ci.uchicago.edu/science-at-beagle/>	
>> Training
>> Machine Learning Fundamentals 
>> Wednesday June 6th, 10AM
>> Room 240A, at the Computation Institute of the University of Chicago
>> 
>> Topics will include:
>> 
>> Nearest neighbor methods and families of distance functions
>> Generative modeling for classification using the multivariate Gaussian
>> Linear regression and its variants
>> Logistic regression
>> Linear classification using the support vector machine
>> Nonlinear modeling using basis expansion and kernel methods
>> Decision trees, boosting, and random forests
>> Methods for flat and hierarchical clustering
>> Principal component analysis
>> Beagle2 Events <http://beagle.ci.uchicago.edu/trainings-and-events/> Learn more about Beagle2 training.
>> Resources
>> Beagle2 Wiki <https://wiki.uchicago.edu/display/Beagle/Welcome+to+Beagle> Get detailed usage information from the Beagle2 team
>> Beagle2 Support <mailto:beagle-support at lists.uchicago.edu> Contact the Beagle2 experts for help
>> Globus <https://www.globusonline.org/> for file transfer. Get started moving files to/from Beagle2 using this fast service
>> Other CI resources <https://www.ci.uchicago.edu/> Learn about other computing resources available at the Computation Institute
>>              
>> Beagle2 Related Publications
>> Manching H, Sengupta S, Hopper KR, Polson SW, Ji Y, Wisser RJ. 
>> Phased Genotyping-by-Sequencing Enhances Analysis of Genetic Diversity and Reveals Divergent Copy Number Variants in Maize 
>> G3 (Bethesda). 2017 Jul 5;7(7):2161-2170. doi: 10.1534/g3.117.042036. PMCID:PMC5499125
>> Wentian Guo, Sue-Jane Wang, Shengjie Yang, Henry Lynn, Yuan Ji 
>> A Bayesian interval dose-finding design addressing Ockham's razor: mTPI-2 
>> Cont. Clinical Trials. July 2017. Vol. 58, Pg. 23–33, DOI: http://dx.doi.org/10.1016/j.cct.2017.04.006 <http://dx.doi.org/10.1016/j.cct.2017.04.006>
>> Rangarajan D. Nadadur, Michael T. Broman, Bastiaan Boukens, et al. 
>> Pitx2 modulates a Tbx5-dependent gene regulatory network to maintain atrial rhythm 
>> Sci Transl Med. 2016 August 31; 8(354): 354ra115. PMCID: PMC5266594
>> C.K. Li, P. Tzeferacos, D. Lamb, G. Gregori et al. 
>> Scaled laboratory experiments explain the kink behaviour of the Crab Nebula jet 
>> Nature Communications, DOI: 10.1038/ncomms13081, 7 Oct 2016
>> Esmael J. Haddadian, Hao Zhang, Karl F. Freed, and Jack F. Douglas 
>> Comparative Study of the Collective Dynamics of Proteins and Inorganic Nanoparticles 
>> Sci Rep. 2017; 7: 41671. doi: 10.1038/srep41671, PMCID: PMC5296861
>> Request for RECENT publications made using Beagle2
>> Please send us your most updated list, including papers that are only under revision. Feel free to add also older ones, since they might not be in our lists yet and press releases that talk about your work with Beagle.
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