The Duel Functions of Acetyl-CoA Carboxylase

Louisiana State University, Baton Rouge LA

Investigators

Abstract

Membranes are the essential barriers that define and delineate the inside versus the outside of a cell. Membranes are composed of fatty acids, and in contrast to humans, bacteria synthesize all of the needed fatty acids for their cellular membranes. Fatty acids are synthesized by a series of enzyme catalyzed chemical reactions. The enzyme that catalyzes the first reaction in fatty acid synthesis is acetyl-CoA carboxylase. Acetyl-CoA carboxylase is a multifunctional enzyme composed of two different enzymes: biotin carboxylase and carboxyltransferase. The first aim of this project involves trying to understand how the three-dimensional structure of biotin carboxylase allows the enzyme to catalyze the reaction. For instance, the enzyme is composed of two identical subunits where each subunit can in theory catalyze the reaction. The fundamental question is: if each subunit is capable of catalyzing the reaction why does it exist as a dimer? One hypothesis that is being tested is that the subunits are not independent but instead depend on each other by alternating their catalytic cycles. Similar to the up and down motion of a bicycle pedal, one subunit catalyzes a reaction while the other releases products. The second and third aims of the project involve trying to understand how the three-dimensional structure of carboxyltransferase participates in regulating the genes that code for the protein. In contrast to biotin carboxylase, carboxyltransferase is composed of two different subunits. The genes coding for these subunits are located at different positions in the bacterial chromosome yet the cell requires equal amounts of each subunit to make a functional enzyme. The fundamental question here is: how does a bacterium regulate expression of the genes coding for the subunits to obtain equal amounts? The carboxyltransferase molecule in bacteria contains a structural motif called a zinc finger that binds to RNA. The hypothesis is that carboxyltransferase regulates expression of both genes by binding to the RNA derived from those genes. When nutrients are low, carboxyltransferase binds to the RNA and inhibits protein synthesis. When nutrients are abundant and new membranes are needed, the substrate for the enzyme acetyl-CoA competes with RNA for binding and fatty acids are synthesized. Thus, carboxyltransferase appears to have two mutually exclusive "duel" functions: catalysis and translational regulation. Broader Impacts of the Research: This project addresses fundamental questions in enzyme catalysis, subunit interactions, and regulation of translation in a model system. The eclectic approach will provide a detailed understanding of structure-function relationships that will have considerable impact on our view of the roles of the enzyme on the physiological level. This project will rely heavily on diverse graduate and undergraduate students to perform the experiments, serving as a training platform for the future scientific workforce using the eclectic tools of biochemistry and molecular biology. In addition, because the enzyme in this project is a carboxylase (using a form of carbon dioxide as substrate) the results of the research may have ramifications for engineering enzymes to reduce the carbon footprint in the environment. Lastly, because membranes are essential for life, acetyl-CoA carboxylase in plants is a logical target for herbicides. Understanding the structure and function of the enzyme can facilitate the design of more potent and environmentally safe herbicides.