Phospholipid Binding and Membrane Attachment of the Osh4 Protein

Abstract

Osh4 is an oxysterol binding protein (OSBP) 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 binding a single sterol residue within a hydrophobic binding pocket and transporting it, against a concentration gradient, from the endoplasmic reticulum to the plasma membrane. Phosphoinositides (PIPs) are thought to stimulate sterol transfer by binding to the Osh4 protein surface. In order to study how the Osh4 protein attaches to the plasma membrane, possible lipid binding sites were investigated through the use of blind docking techniques. Model ligand compounds for phosphatidylcholine, phosphatidylserine, and two PIP [PI(4,5)P2 and PI(3,4,5)P3] head groups were docked against several conformational snapshots of the Osh4 surface to determine possible regions favorable to interact with plasma membrane lipids. These conformational snapshots were taken from two 25-ns molecular dynamics simulations of the Osh4 protein complexed with ergosterol and two complexed with 25-hydroxycholesterol. The PIP models frequently docked to a lysine-rich region on an exposed portion of the protein's β-barrel. This region is bounded by a flexible surface loop that is believed to be important for Osh4-membrane binding. Osh4-membrane interaction was also investigated through molecular dynamics simulations of a combined membrane and protein system. Model sterol-acceptor and sterol-donor membranes were constructed using CHARMM-GUI and equilibrated for 25 ns. Key residues of the Osh4 protein that were identified during the blind docking tests, as well as residues known to be in close contact with these membranes upon binding, were placed parallel to the membrane surface. Ultimately, understanding how Osh4 attaches to cellular membranes will lead to a clear understanding how this protein transports sterols in vivo.