TCBG Seminar

Flexible Architecture of Ca2+ Release Channels by Cryo-EM

Professor Irina I. Serysheva
The University of Texas Medical School at Houston
Houston, TX

Monday, March 14, 2011
3:00 pm (CT)
3269 Beckman Institute


Ca2+ release channels are essential to a wide variety of cellular processes, including muscle contraction, heartbeat and brain function. They are targets for many drugs used to treat numerous diseases that are associated with abnormal regulation of cell Ca2+ levels. Our research focuses on two members of an intracellular Ca2+ release channel family: type-1 ryanodine receptor (RyR1) and type-1 inositol 1,4,5-trisphosphate receptor (IP3R1). Localized in ER/SR membranes, these channels form large tetrameric protein complexes with a molecular mass of ~2.3 MDa for RyR1 and ~1.3 MDa for IP3R1. The RyR1 is the primary Ca2+ release channel in skeletal muscle. The IP3R1 channel is detected in virtually all cell types with highest densities in the cerebellum. Ca2+ release channels function as scaffold membrane proteins, comprising of multiple interaction and regulatory motifs with distinctive features enabling them to convert diverse cellular signals into Ca2+ signals. The structural analysis of these channels has been hampered by both their enormous size and interaction with lipid membranes in their native state, making X-ray or NMR techniques poorly suited for structural studies of these membrane protein complexes. Single particle electron cryo-microscopy (cryo-EM) emerges as a powerful technique allowing researchers to study not only this kind of macromolecular protein complexes, but also to choose conditions favoring a particular functional state of the protein of interest. Using single particle cryo-EM, we have determined structure of both RyR1 and IP3R1 channels at ~1-nm resolution in the closed state. These cryo-EM density maps clearly resolved the 3D molecular organization of the channel proteins, including molecular boundaries between the monomeric subunits, and the secondary structure elements in both the TM and CY regions. In the TM region, we observe a twisted bundle of four α- helices that form a funnel shaped structure around the 4-fold symmetry axis, strikingly similar to the central ion-conduction pore of K+ channels. While the TM region of the IP3R1 channel is quite similar to RyR1 and other ion channels, the CY region has a very different architecture. 3D statistical analysis of the cryo-EM density map identifies high variance in the CY region of IP3R1. This structural variation could be attributed to genuine structural flexibility of Ca2+ release channels, and/or the presence of unknown modulators co-purified with the channel protein, subtly altering its conformation.

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