Binding of Isoflurane to Glic Alters the Structure and Dynamics of the Protein

Abstract

General anesthetics are thought to interact with proteins to produce anesthesia. Most of the experimental efforts to date have been focused on searching for discrete anesthetic binding sites. How exactly anesthetics work after the events of anesthetic binding is rarely explored. With the recent availability of two crystal structures for the bacterial Gloeobacter violaceus pentameric ligand-gated ion channel (GLIC), which is sensitive to a variety of general anesthetics, we performed multiple replica molecular dynamics simulations for more than 100 ns with and without the general anesthetic isoflurane. Multiple sites within the protein, primarily in the extracellular domain and at the interface of extracellular and transmembrane domains, provided favorable binding environments for isoflurane, showing little isoflurane translational displacement over the course of the simulations. Isoflurane caused changes to the structure and dynamics of GLIC when compared to the control simulation. Subunit-subunit interactions in the extracellular domain of the pentamer were disrupted, resulting in increased flexibility of the subunit pairs, as measured by Gaussian network model analysis. Salt-bridge linkages between and within the extracellular region of subunits containing isoflurane binding sites were altered by the presence of isoflurane, reflecting a change in the tertiary and quaternary structure of the channel. Changes were not isolated to the extracellular domain. The subunits containing more isoflurane binding sites also showed a greater change in tilt angle of the second transmembrane helices. Taken together, anesthetic binding at subunit interfaces can lead to changes in quaternary structure and global dynamics, causing allosteric changes in channel motional characteristics critical to gating. Supported by the NIH (R01GM56257, R01GM66358, R37GM049202, and T32GM075770), and the NSF through TeraGrid resources provided by PSC, NICS and TACC (TG-MCB050030N).