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bioRxiv · 10.1101/2021.01.18.426261

Polymeric assembly of endogenous Tuberous Sclerosis Protein Complex

Dai, D. L.·Hasan, S. M. N.·Woollard, G.·Bueler, S. A.·Julien, J.-P.·Rubinstein, J. L.·Mazhab-Jafari, M. T.
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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 regulation of growth and energy consumption. Biochemical and electron cryomicroscopy (cryoEM) studies of recombinant pTSC have revealed the structure and stoichiometry of the pTSC and have hinted at the possibility that the complex 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 up to 1 m in length in linear and web-like organizations prepared from pig brain. These observations suggest polymerization of endogenous pTSC into filamentous super-structures.

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Dai, D. L., Hasan, S. M. N., Woollard, G., Bueler, S. A., Julien, J.-P., Rubinstein, J. L., Mazhab-Jafari, M. T.. 2021-01-19. Polymeric assembly of endogenous Tuberous Sclerosis Protein Complex. https://doi.org/10.1101/2021.01.18.426261

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