Please use this identifier to cite or link to this item:
Title: Purification of Integral Membrane Proteins and Lipid-Binding Assays
Authors: Gross, D.A.
Silver, D.L. 
Keywords: Adipocytes
Detergent micelles
Lipid droplets
Issue Date: 2013
Citation: Gross, D.A., Silver, D.L. (2013). Purification of Integral Membrane Proteins and Lipid-Binding Assays. Methods in Cell Biology 116 : 191-211. ScholarBank@NUS Repository.
Abstract: The lipid droplet (LD) is an evolutionarily conserved organelle composed primarily of triglycerides (TAG) and cholesteryl esters. Recently, Fat storage-. Inducing Transmembrane proteins 1 & 2 (FITM1/FIT1 and FITM2/FIT2) were discovered as a conserved family of proteins involved in fat storage. FIT1 and FIT2 are both localized to the endoplasmic reticulum, but have distinct tissue distributions. FIT proteins mediate TAG LD accumulation when overexpressed, but do not synthesize TAG. FIT proteins function by partitioning newly synthesized TAG into LDs. In order to understand the mechanism by which this occurs, a method was developed to purify FIT proteins from insect cells in detergent micelles. The ability of purified FIT proteins to bind TAG and other neutral lipids was tested in detergent micelles, demonstrating lipid specificity and saturation binding. These techniques can be applied to a variety of proteins in lipid biology in an effort to try to reconstitute a mechanism of action or protein activity. The methods that will be discussed here can also be scaled to either screen a library of mutant proteins for binding to a particular compound or utilized to delineate structural requirements of ligands that are important for protein-ligand interactions. Here, we present a detailed description of the purification protocol and micellar protein-ligand binding experiments and their possible applications. © 2013 Elsevier Inc.
Source Title: Methods in Cell Biology
ISSN: 0091679X
DOI: 10.1016/B978-0-12-408051-5.00010-3
Appears in Collections:Staff Publications

Show full item record
Files in This Item:
There are no files associated with this item.

Google ScholarTM



Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.