Belmont University Research Symposium (BURS)

Optimizing Purification of Gle1 for Future Analysis of Dbp5 ATPase activity

Publication Date

2024

College

Sciences and Mathematics, College of

Department

Chemistry and Physics, Department of

BURS Faculty Advisor

Dr. Rigsby and Dr. Adams

Presentation Type

Oral Presentation

Abstract

The mRNA nuclear export process is a tightly regulated biological system important for eukaryotic gene expression. Gene expression is of paramount importance in eukaryotes and prokaryotes alike as it governs the process through which genetic information encoded in DNA can produce functional products, such as proteins. This central dogma of life orchestrates complex molecular processes that underpin how biology is understood, such as basic cellular functions universal to all living organisms. The export process begins with mRNA sequestered inside the nucleus, where the transcript associates with the protein, Mex67. The duplex gets transported through nuclear pore complexes (NPCs) out of the nucleus and into the cytoplasm, the site of translation. In the cytoplasm, Mex67 is removed from the transcript, which is facilitated by the binding of the protein complex Dbp5-ATP, Gle1, IP6, and Nup42. Gle1 stimulates ATPase activity of Dbp5, which enables binding to the transcript and catalyzes the hydrolysis from ATP to ADP, ultimately releasing mRNA for the proteins to be recycled. Therefore, knowing the extent at which Gle1 stimulates Dbp5 ATPase activity allows for a better understanding of the mRNA export mechanism, which could lead to preventing harmful changes in humans, such as Lethal Congenital Contracture Syndrome 1 (LCCS1). Throughout this study, a biochemical system was employed to ensure optimized induction of Gle1 in E. coli Rosetta cells. Other efforts of the system included sonicating the induced cells for successful extraction of Gle1. In future research, the extracted Gle1 will be purified for in vitro analysis to determine its factors influencing Dbp5 activity.

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