Structural and Mechanistic Insights into Regulation of RGS17 and PLCƐ

Monita Sieng, Purdue University

Abstract

Recent advances in structural biology and biochemistry have identified proteins downstream of G protein-coupled receptors (GPCRs) as promising drug targets. These proteins are highly regulated to ensure proper physiological responses from extracellular stimuli. Dysregulation of these signaling enzymes can have detrimental consequences, including cardiovascular disease and cancer. Understanding how these proteins are regulated from a structural and biochemical standpoint can therefore be exploited to develop new therapeutics. In this work, the molecular mechanism of regulation of two different proteins downstream of GPCRs is investigated. The first protein, Regulator of G Protein Signaling 17 (RGS17), is involved in numerous processes throughout the body, including the development and progression of lung cancer. This work presents the crystal structure of RGS17 bound to Ca2+. Ca2+ was found to bind to the same site as the predicted Gα binding surface and increases interactions between RGS17 and Gαo. Therefore, Ca2+ positively regulates RGS17, supporting a mechanism in which Ca2+ increases the GTPase activating function of the RZ-family of RGS proteins to ultimately downregulate Ca2+signaling. The second protein, phospholipase CƐ(PLCƐ), has been implicated in cardiac hypertrophy through its production of second messengers. This process is regulated by the small GTPase Rap1A. This work presents insight into the molecular mechanism of Rap1A-dependent activation of PLCƐ, in which four conserved, hydrophobic residues on the surface of the RA2 domain of PLCƐ play an essential role. Furthermore, small angle X-ray scattering studies show that binding of Rap1A induces conformational changes in PLCƐ, resulting in a more compact activated complex. This supports a mechanism in which Rap1A is an allosteric activator of PLCƐ, inducing conformational changes in PLCƐthat increase lipid hydrolysis by relieving autoinhibitory interactions and/or by promoting interactions with the cell membrane.

Degree

Ph.D.

Advisors

Lyon, Purdue University.

Subject Area

Biochemistry|Physiology|Medical imaging|Medicine|Neurosciences|Oncology|Pharmaceutical sciences|Polymer chemistry|Public health

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