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The mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) is a master regulator of metabolism and cell growth. Among the numerous extracellular and intracellular signals, certain amino acids activate mTORC1 in a Rag-dependent manner. Arginine can stimulate mTORC1 activity by releasing the inhibitor CASTOR1 (Cellular Arginine Sensor of mTORC1) from GATOR2, a positive regulator of mTORC1 which interacts with GATOR1, the GAP for RagA/B. Three groups have resolved the structures of arginine-CASTOR1 complex, shedding a new light on molecular basis of the regulation of mTORC1 activity by arginine. However, lacking the apo structure of CASTOR1 prelimited the molecular understanding of mechanism underlying mTORC1 regulation. Here, we report crystal structures of arginine sensor CASTOR1 in arginine-bound and ligand free states at 2.05 Å and 2.8 Å, respectively. Structural comparison of CASTOR1 between two states reveals near identical conformations, except in two loop regions. It indicates CASTOR1 does not undergo large conformational change during arginine binding. Therefore, we conclude a detailed structural interpretation of arginine sensing by CASTOR1 in mTORC1 pathway., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The major facilitator superfamily (MFS) is the largest secondary transporter family and is responsible for transporting a broad range of substrates across the biomembrane. These proteins are involved in a series of conformational changes during substrate transport. To decipher the transport mechanism, it is necessary to obtain structures of these different conformations. At present, great progress has been made in predicting protein structure based on coevolutionary information. In this study, AlphaFold2 was used to predict different conformational structures for 69 MFS transporters of E. coli after the selective mutation of residues at the interface between the N- and C-terminal domains. The predicted structures for these mutants had small RMSD values when compared to structures obtained using X-ray crystallography, which indicates that AlphaFold2 predicts the structure of MSF transporters with high accuracy. In addition, different conformations of other transporter family proteins have been successfully predicted based on mutation methods. This study provides a structural basis to study the transporting mechanism of the MFS transporters and a method to probe dynamic conformation changes of transporter family proteins when performing their function. [ABSTRACT FROM AUTHOR]