Using lipid nanodiscs for the in vitro glycosylation of membrane proteins

Abstract

Membrane proteins are involved in a number of human diseases are important to study. Due to the fact that these proteins are, by nature, insoluble in aqueous solutions, they require detergent or lipid environments that resemble cell membranes to remain soluble. Nanodiscs can be used to prevent these proteins from precipitating. Nanodiscs are patches of phospholipid bilayer that is encircled by membrane scaffold protein and mimic the thickness of the human cell membrane. This allows for membrane proteins to be studied in a native-like environment. In our lab we study a specific group of proteins that are post-translationally modified. These proteins are glycoproteins which have sugar groups attached to their sidechains. It is also important to understand how proteins change when a sugar is attached to them. This will be examined by first inserting the membrane protein into nanodiscs and then attempting to glycosylate them using in vitro glycosylation with the enzyme N-Glycosyltransferase (NGT). We will show that membrane proteins that are inserted into nanodiscs can be glycosylated using these methods.