| Products/Services Used | Details | Operation |
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| Protein and Antibody Isolation | glycerol, 10 mM NaF, and 2 mM dithiothreitol (DTT). Lysate was clarified by centrifugation, filtered at 0.8 m, and then incubated with anti-FLAG resin (GenScript, L00432-25) for 30 min at 4 C with agitation. The resin was washed with lysis buffer, and TSC was eluted with lysis buffer containing FLAG peptide (0. for 30 min at 4 C with agitation. The resin was washed with lysis buffer, and TSC was eluted with lysis buffer containing FLAG peptide (0.1 mg ml 1 ; GenScript). TSC was further purified by anion-exchange chromatography using a Mono Q 5/50 GL (Cytiva) with a linear gradient of 100 to 1000 mM NaCl in 20 mM Hepes | Get A Quote |
Tuberous sclerosis complex (TSC) is targeted to the lysosomal membrane, where it hydrolyzes RAS homolog mTORC1 binding (RHEB) from its GTP-bound to GDP-bound state, inhibiting mechanistic target of rapamycin complex 1 (mTORC1). Loss-of-function mutations in TSC cause TSC disease, marked by excessive tumor growth. Here, we overcome a high degree of continuous conformational heterogeneity to determine the 2.8- cryo electron microscopy (cryo-EM) structure of the complete human TSC in complex with the lysosomal recruitment factor WD repeat domain phosphoinositide interacting protein 3 (WIPI3). We discover a previously undetected amino-terminal TSC1 HEAT repeat dimer that clamps onto a single TSC wing and forms a ph... More
