Plasma Membrane Topology of the Insulin Receptor: Insights from Computational Modeling

Abstract

The insulin receptor in the plasma membrane is composed of two alpha-subunits, each linked to a beta-subunit and to each other by disulfide bonds. It has been proposed that both alpha-subunits are extracellular and anchored to the plasma membrane via a single transmembrane (TM) helix in the N-terminal region of each beta-subunit. However, we find that newer sequence analysis algorithms predict that the alpha-subunits of human insulin receptor (P06213) are largely intracellular and each contains a single TM helix in the N-terminal region. The MEMSAT-SVM method predicts (after correcting for a 19 residue N-terminal signal peptide) that the alpha-subunit contains a TM helix between residues 64 and 79, with a 63 amino acid ectodomain and a large 676 residue cytoplasmic domain. The existence of a large intracellular domain is consistent with the presence of 10 caveolin-binding (CB) motifs (OxOxxxxO, OxxxxOxxO, where O is the aromatic amino acid F, Y, or W), which exist within most caveolin-associated proteins where caveolin is accessible only intracellularly. Our computational analysis of the insulin receptor beta-subunit sequence confirms a single TM helix between residues 194 - 218 (UniProtKB places the TM helix within residues 194-213), with 1 CB motif proximal to the TM helix (W175 - Y182). Similar topologies were obtained with insulin receptor sequences from a variety of species. The insulin receptor co-isolates with lipids and the purified insulin receptor readily forms lipid complexes. Thus, caveolin binding motifs may direct the insulin receptor tetramer to caveolin-rich lipid rafts. The topology of the insulin receptor and CB motifs is relevant to understanding the microdynamics of insulin receptor movement between caveolar and non-caveolar regions of the plasma membrane as well as insulin-mediated events such as endocytosis, membrane hyperpolarization, and regulation of Na+/K+/H+ ions.