Binding of A Natural Sterol to the Osh4 Protein of Yeast and Membrane Attachment

Osh4 is an oxysterol binding protein homologue found in yeast that is essential for the intracellular transport of sterols and for cell life. It has been proposed that Osh4 acts as a lipid transport protein, capable of carrying sterols from the endoplasmic reticulum to the plasma membrane (PM).Molecular dynamics (MD) simulations were used to analyze the binding of ergosterol to the Osh4 protein on an atomic level. During the course of 25-ns simulations, the sterol molecule remained tightly bound to the binding pocket of Osh4. These simulations revealed ergosterol binding was aided by both water-mediated interactions between the 3-hydroxyl (3-OH) group of ergosterol and surrounding polar residues as well as direct hydrogen binding between ergosterol and the Trp46 and Gln96 residues. Analysis of the interaction energy between ergosterol and Gln96 shows distinctly different states (−9.3 kcal/mol, −7.3 kcal/mol, and −4 kcal/mol), where the highest energy state was encountered 77% of the time.In order to study how the Osh4 protein attaches to the PM, possible lipid binding sites were investigated through the use of a docking program as well as MD simulations. A model compound consisting of a phosphocholine lipid head group truncated at the C2 carbon solvated the Osh4 protein in conjunction with water. Protein/lipid interactions were observed and used to determine the proper orientation and placement of the protein with respect to a model membrane. We also developed a model yeast membrane containing ergosterol using CHARMM-GUI, and key membrane properties were investigated using data collected from a 60-ns MD simulation (areas per lipid, density profiles, and lipid dynamics). Ultimately, understanding how Osh4 attaches to the PM will lead to a clear understanding on how this protein transports sterols in vivo.