CAREER: Biochemical and Structural Mechanisms Controlling tRNA-Modifying Metalloenzymes

University Of Delaware, Newark DE

Investigators

Abstract

With the support of the Chemistry of Life Processes (CLP) program in the Division of Chemistry (CHE) and the Established Program to Stimulate Competitive Research (EPSCoR), Jeffrey Mugridge from the University of Delaware is studying the biochemical and structural mechanisms controlling metalloenzymes that install important chemical modifications on transfer RNA (tRNA) molecules. Cellular tRNA molecules are decorated with a huge diversity of chemical modifications that are essential for correctly tuning tRNA structure, stability, and efficient and accurate protein synthesis. Defects in the enzymes and biochemical pathways that install tRNA modifications are linked to a wide range of human diseases from cancers to neurodegenerative disorders. However, for many of the enzymes that install key chemical modifications on tRNA, a clear understanding of how these enzymes selectively carry out their modification reactions is missing. Filling this gap in knowledge is important for our fundamental understanding of cellular RNA biology. This proposal will define the detailed atomic-level mechanisms for two distinct classes of metal-dependent enzymes (metalloenzymes) that install key modifications on tRNA that directly impact protein synthesis. Closely integrated with this research, a course-based undergraduate research experience (CURE) will be developed that engages both undergraduates and high-school students from underrepresented groups in a collaborative, discovery-based course where students will make and experimentally test predictions about protein-tRNA interactions. The outcomes from this work could provide new information on how tRNA-modifying metalloenzymes carry out complex, multistep reactions on tRNA and broaden engagement of students in research-focused activities at the undergraduate and high-school levels. Hypermodification of the tRNA anticodon loop is essential for proper codon-anticodon recognition and mRNA decoding in the ribosome during translation. The cell installs numerous, chemically complex modifications at these locations to ensure and control translational efficiency and fidelity. This project combines techniques from biochemistry, structural biology, biophysics, and chemical biology to study two different classes of tRNA-modifying metalloenzymes that install sequential modifications on the anticodon loop of tRNA. The goals of this work are to: (1) define the stepwise chemical mechanisms used by these classes of metalloenzymes to install anticodon loop modifications, (2) unveil the structural mechanisms and protein conformations that control tRNA recognition and modification selectivity, and (3) provide new, broad information about how these classes of metalloenzymes are regulated in the cell. The research outcomes could deepen our basic understanding of tRNA biology and pave the way for future therapeutic development targeting these or similar metalloenzymes and tRNA modification pathways. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.