Detergent-mediated protein aggregation.

Because detergents are commonly used to solvate membrane proteins for structural evaluation, much attention has been devoted to assessing the conformational bias imparted by detergent micelles in comparison to the native environment of the lipid bilayer. Here, we conduct six 500-ns simulations of a system with >600,000 atoms to investigate the spontaneous self assembly of dodecylphosphocholine detergent around multiple molecules of the integral membrane protein PagP. This detergent formed equatorial micelles in which acyl chains surround the protein's hydrophobic belt, confirming existing models of the detergent solvation of membrane proteins.

Assessing the stability of membrane proteins to detect ligand binding using differential static light scattering.

Protein stabilization upon ligand binding has frequently been used to identify ligands for soluble proteins. Methods such as differential scanning fluorimetry (DSF) and differential static light scattering (DSLS) have been employed in the 384-well format and have been useful in identifying ligands that promote crystallization and 3D structure determination of proteins. However, finding a generic method that is applicable to membrane proteins has been a challenge as the high hydrophobicity of membrane proteins and the presence of detergents essential for their solubilization interfere with fluorescence-based detections.

Biochemical characterization of MsbA from Pseudomonas aeruginosa.

Lipopolysaccharide of Pseudomonas aeruginosa is a major constituent of the outer membrane, and it is composed of three distinct regions: lipid A, core oligosaccharide, and O antigen. Lipid A and core oligosaccharides (OS) are synthesized and assembled at the cytoplasmic side of the inner membrane and then translocated to the periplasmic side of the membrane where lipid A-core becomes the acceptor of the O antigens. Here we show that MsbA encoded by pA4997 of the P.