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Structural Characterization of Endogenous Tuberous Sclerosis Protein Complex Revealed Potential Polymeric Assembly.
- Source :
-
Biochemistry [Biochemistry] 2021 Jun 15; Vol. 60 (23), pp. 1808-1821. Date of Electronic Publication: 2021 Jun 03. - Publication Year :
- 2021
-
Abstract
- 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 (F <subscript>ab</subscript> ) specific for the TSC2 subunit of pTSC. We demonstrate F <subscript>ab</subscript> -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.
- Subjects :
- Animals
Cryoelectron Microscopy methods
Cytoskeleton metabolism
Humans
Protein Binding physiology
Rats
Recombinant Proteins metabolism
Signal Transduction genetics
Swine
Tuberous Sclerosis genetics
Tuberous Sclerosis Complex 2 Protein metabolism
Tumor Suppressor Proteins metabolism
Tuberous Sclerosis metabolism
Tuberous Sclerosis Complex 2 Protein chemistry
Tuberous Sclerosis Complex 2 Protein ultrastructure
Subjects
Details
- Language :
- English
- ISSN :
- 1520-4995
- Volume :
- 60
- Issue :
- 23
- Database :
- MEDLINE
- Journal :
- Biochemistry
- Publication Type :
- Academic Journal
- Accession number :
- 34080844
- Full Text :
- https://doi.org/10.1021/acs.biochem.1c00269