TR&D Project 3. The Analysis Stage II: Tools for Analyzing the Connectivity and Morphology of Macromolecular Assemblies As the structure of an assembly is directly related to its functional role, we aim to define the architecture of isolated native macromolecular complexes of interest. For this, our molecular microscope pipeline needs information on the shape and connectivity of an assembly?s components, accurately representing their arrangement at the highest spatial and temporal resolution. Thus, we will develop and refine methodologies to determine the morphologies and spatial relationships between components within complexes at several distance scales, ranging from a description of the overall subunit shape and arrangement to defining atomic resolution contacts between pairs of macromolecules. Our strategy entails using orthologous methods, in order to provide complementary data and to cover a wide range of resolutions, to inform us about the shape, dimensions and connectivity of single proteins and macromolecular assemblies. We will focus on methods that have already proven particularly empowering, but which have significant scope for further advancement. These include electron microscopy (EM) and chemical cross-linking with mass spectrometry (XL-MS): through the former, we can produce morphological maps with sufficient detail to resolve the shapes and locations of complexes, proteins, domains and folds; in parallel, through the latter, we will obtain information on how each component of the assembly is positioned relative to all other components, and the entire structure. These data when combined with data from complementary, well-established methods, will be used to generate structural models of assemblies. To elucidate cellular functions, we propose to gather and interpret dynamic data about the changing morphologies of assemblies, and the changing interactions within these assemblies. Our molecular microscope pipeline thus seeks to build concrete high precision 3D models, and 4D models that change in time.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Biotechnology Resource Grants (P41)
Project #
5P41GM109824-07
Application #
9922922
Study Section
Special Emphasis Panel (ZRG1)
Project Start
Project End
Budget Start
2020-05-01
Budget End
2021-04-30
Support Year
7
Fiscal Year
2020
Total Cost
Indirect Cost
Name
Rockefeller University
Department
Type
DUNS #
071037113
City
New York
State
NY
Country
United States
Zip Code
10065
Holden, Jennifer M; Koreny, Ludek; Obado, Samson et al. (2018) Involvement in surface antigen expression by a moonlighting FG-repeat nucleoporin in trypanosomes. Mol Biol Cell 29:1100-1110
Yoshizawa, Takuya; Ali, Rustam; Jiou, Jenny et al. (2018) Nuclear Import Receptor Inhibits Phase Separation of FUS through Binding to Multiple Sites. Cell 173:693-705.e22
Taylor, Martin S; Altukhov, Ilya; Molloy, Kelly R et al. (2018) Dissection of affinity captured LINE-1 macromolecular complexes. Elife 7:
Chan, Ho Lam; Beckedorff, Felipe; Zhang, Yusheng et al. (2018) Polycomb complexes associate with enhancers and promote oncogenic transcriptional programs in cancer through multiple mechanisms. Nat Commun 9:3377
Zinoviev, Alexandra; Goyal, Akanksha; Jindal, Supriya et al. (2018) Functions of unconventional mammalian translational GTPases GTPBP1 and GTPBP2. Genes Dev 32:1226-1241
Schrank, Benjamin R; Aparicio, Tomas; Li, Yinyin et al. (2018) Nuclear ARP2/3 drives DNA break clustering for homology-directed repair. Nature 559:61-66
Hua, Nan; Tjong, Harianto; Shin, Hanjun et al. (2018) Producing genome structure populations with the dynamic and automated PGS software. Nat Protoc 13:915-926
Hayama, Ryo; Sparks, Samuel; Hecht, Lee M et al. (2018) Thermodynamic characterization of the multivalent interactions underlying rapid and selective translocation through the nuclear pore complex. J Biol Chem 293:4555-4563
Guy, Andrew J; Irani, Vashti; Beeson, James G et al. (2018) Proteome-wide mapping of immune features onto Plasmodium protein three-dimensional structures. Sci Rep 8:4355
Sparks, Samuel; Temel, Deniz B; Rout, Michael P et al. (2018) Deciphering the ""Fuzzy"" Interaction of FG Nucleoporins and Transport Factors Using Small-Angle Neutron Scattering. Structure 26:477-484.e4

Showing the most recent 10 out of 111 publications