Binding and release of cholesterol in the Osh4 protein of yeast

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Abstract
Sterols have been shown experimentally to bind to the Osh4 protein (a homolog of the oxysterol binding proteins) of Saccharomyces cerevisiae within a binding tunnel, which consists of antiparallel β‐sheets that resemble a β‐barrel and three α‐helices of the N‐terminus. This and other Osh proteins are essential for intracellular transport of sterols and ultimately cell life. Molecular dynamics (MD) simulations are used to study the binding of cholesterol to Osh4 at the atomic level. The structure of the protein is stable during the course of all MD simulations and has little deviation from the experimental crystal structure. The conformational stability of cholesterol within the binding tunnel is aided in part by direct or water‐mediated interactions between the 3‐hydroxyl (3‐OH) group of cholesterol and Trp46, Gln96, Tyr97, Asn165, and/or Gln181 as well as dispersive interactions with Phe42, Leu24, Leu39, Ile167, and Ile203. These residues along with other nonpolar residues in the binding tunnel and lid contribute nearly 75% to the total binding energy. The strongest and most populated interaction is between Gln96 and 3‐OH with a cholesterol/Gln96 interaction energy of −4.5 ± 1.0 kcal/mol. Phe42 has a similar level of attraction to cholesterol with −4.1 ± 0.3 kcal/mol. A MD simulation without the N‐terminus lid that covers the binding tunnel resulted in similar binding conformations and binding energies when compared with simulations with the full‐length protein. Steered MD was used to determine details of the mechanism used by Osh4 to release cholesterol to the cytoplasm. Phe42, Gln96, Asn165, Gln181, Pro211, and Ile206 are found to direct the cholesterol as it exits the binding tunnel as well as Lys109. The mechanism of sterol release is conceptualized as a molecular ladder with the rungs being amino acids or water‐mediated amino acids that interact with 3‐OH. Proteins 2009. Published 2008 Wiley‐Liss, Inc.