Project description:The insulin receptor is a dimeric protein that has a crucial role in controlling glucose homeostasis, regulating lipid, protein and carbohydrate metabolism, and modulating brain neurotransmitter levels. Insulin receptor dysfunction has been associated with many diseases, including diabetes, cancer and Alzheimer's disease. The primary sequence of the receptor has been known since the 1980s, and is composed of an extracellular portion (the ectodomain, ECD), a single transmembrane helix and an intracellular tyrosine kinase domain. Binding of insulin to the dimeric ECD triggers auto-phosphorylation of the tyrosine kinase domain and subsequent activation of downstream signalling molecules. Biochemical and mutagenesis data have identified two putative insulin-binding sites, S1 and S2. The structures of insulin bound to an ECD fragment containing S1 and of the apo ectodomain have previously been reported, but details of insulin binding to the full receptor and the signal propagation mechanism are still not understood. Here we report single-particle cryo-electron microscopy reconstructions of the 1:2 (4.3?Å) and 1:1 (7.4?Å) complexes of the insulin receptor ECD dimer with insulin. The symmetrical 4.3?Å structure shows two insulin molecules per dimer, each bound between the leucine-rich subdomain L1 of one monomer and the first fibronectin-like domain (FnIII-1) of the other monomer, and making extensive interactions with the ?-subunit C-terminal helix (?-CT helix). The 7.4?Å structure has only one similarly bound insulin per receptor dimer. The structures confirm the binding interactions at S1 and define the full S2 binding site. These insulin receptor states suggest that recruitment of the ?-CT helix upon binding of the first insulin changes the relative subdomain orientations and triggers downstream signal propagation.

| S-EPMC5886813 | biostudies-literature

Single-particle cryo-EM structural studies of the β<sub>2</sub>AR-Gs complex bound with a full agonist formoterol.

Project description: Not available

| S-EPMC7338445 | biostudies-literature

Cryo-EM single-particle structure refinement and map calculation using Servalcat.

Project description:In 2020, cryo-EM single-particle analysis achieved true atomic resolution thanks to technological developments in hardware and software. The number of high-resolution reconstructions continues to grow, increasing the importance of the accurate determination of atomic coordinates. Here, a new Python package and program called Servalcat is presented that is designed to facilitate atomic model refinement. Servalcat implements a refinement pipeline using the program REFMAC5 from the CCP4 package. After the refinement, Servalcat calculates a weighted Fo - Fc difference map, which is derived from Bayesian statistics. This map helps manual and automatic model building in real space, as is common practice in crystallography. The Fo - Fc map helps in the visualization of weak features including hydrogen densities. Although hydrogen densities are weak, they are stronger than in the electron-density maps produced by X-ray crystallography, and some H atoms are even visible at ∼1.8 Å resolution. Servalcat also facilitates atomic model refinement under symmetry constraints. If point-group symmetry has been applied to the map during reconstruction, the asymmetric unit model is refined with the appropriate symmetry constraints.

| S-EPMC8489229 | biostudies-literature

Single particle cryo-EM structure of RIG-I bound to the end of p3SLR30 (+AMPPNP)

Project description: Not available

| EMPIAR-11337 | biostudies-other