Single-particle cryo-EM of the ryanodine receptor from skeletal muscle: reconstitution into nanodiscs and the conformational effects of magnesium.

Author

Alex H. Will, Virginia Commonwealth UniversityFollow

DOI

https://doi.org/10.25772/YN0P-YN86

Defense Date

2017

Document Type

Thesis

Degree Name

Master of Science

Department

Physiology and Biophysics

First Advisor

Montserrat Samso

Second Advisor

Jose Miguel Eltit

Third Advisor

John Bigbee

Abstract

The RyR1 acts as an intracellular calcium channel that allows passage of Ca²⁺ from the sarcoplasmic reticulum to the cytoplasm; this increase in cytosolic Ca²⁺ is required for excitation-contraction coupling. Certain mutations in RyR1 have been directly linked to malignant hyperthermia (MH) and central core disease. In a series of studies, the role of Mg²⁺ has been explored as it pertains to MH, and it has been determined that dysregulation of Mg²⁺ can even lead to MH in patients without mutations. Consequently, the aim of the study was to insert the RyR1 into nanodiscs, small, circular lipid bilayers used to solubilize membrane proteins, and to use cryo-electron microscopy to assess the conformation of the RyR1 in the presence of Mg²⁺ and AMP-PCP. Particle reconstruction generated a 9.0 Å resolution map that confirmed successful incorporation of the RyR1 into nanodiscs and allowed visualization of the RyR1 in a physiological closed state.

Rights

© The Author

Is Part Of

VCU University Archives

Is Part Of

VCU Theses and Dissertations

Date of Submission

7-7-1992