Structural Characterization of Endogenous Tuberous Sclerosis Protein Complex Revealed Potential Polymeric Assembly
| Autor: | Yazan M. Abbas, David Dai, Stephanie A. Bueler, Mohammad T. Mazhab-Jafari, Jean-Philippe Julien, S. M. Naimul Hasan, John L. Rubinstein, Geoffrey Woollard |
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| Rok vydání: | 2021 |
| Předmět: |
Swine
Protein subunit Endogeny Biochemistry law.invention 03 medical and health sciences Tuberous Sclerosis law Tuberous Sclerosis Complex 2 Protein Animals Humans Cytoskeleton 0303 health sciences Chemistry Tumor Suppressor Proteins Cryoelectron Microscopy 030302 biochemistry & molecular biology Immunogold labelling Negative stain Recombinant Proteins Rats Tuberous sclerosis protein Recombinant DNA Biophysics TSC2 Electron microscope Protein Binding Signal Transduction |
| Zdroj: | Biochemistry. 60:1808-1821 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/acs.biochem.1c00269 |
| Popis: | Tuberous sclerosis protein complex (pTSC) nucleates a proteinaceous signaling hub that integrates information about the internal and external energy status of the cell in the regulation of growth and energy consumption. Biochemical and cryo-electron microscopy studies of recombinant pTSC have revealed its structure and stoichiometry and hinted at the possibility that the complex may form large oligomers. Here, we have partially purified endogenous pTSC from fasted mammalian brains of rat and pig by leveraging a recombinant antigen binding fragment (Fab) specific for the TSC2 subunit of pTSC. We demonstrate Fab-dependent purification of pTSC from membrane-solubilized fractions of the brain homogenates. Negative stain electron microscopy of the samples purified from pig brain demonstrates rod-shaped protein particles with a width of 10 nm, a variable length as small as 40 nm, and a high degree of conformational flexibility. Larger filaments are evident with a similar 10 nm width and a ≤1 μm length in linear and weblike organizations prepared from pig brain. Immunogold labeling experiments demonstrate linear aggregates of pTSC purified from mammalian brains. These observations suggest polymerization of endogenous pTSC into filamentous superstructures. |
| Databáze: | OpenAIRE |
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