Structural Determinants of Cholesterol Recognition in Helical Membrane Proteins

Abstract

Cholesterol is an integral component of mammalian membranes. It has been shown to modulate membrane dynamics and alter protein function. However, understanding the molecular mechanisms of how cholesterol alters protein function through specific interactions is complicated by our inability to differentiate cholesterol as a ligand and as a lipid in the membrane. Specifically, it is difficult to discern if a cholesterol-membrane protein interaction observed in an experimental structure is specific, unspecific, or a crystallization artefact. The only widely recognized search algorithm for cholesterol binding pockets is sequence-based, i.e. the CRAC and CARC motifs. While, both motifs have been identified on multiple proteins, there is inconclusive evidence on the necessity and sufficiency of its role in identifying cholesterol functional modifiers on its own. To elucidate cholesterol's role on the functional changes of an array of protein types, we characterized the rate of evolution of interacting residues, binding pocket residue types, and binding pocket spatial arrangement of high-resolution crystal structures of cholesterol-membrane protein complexes in the protein data bank (PDB). We compare the predictive power of this multi-layer analysis to the outcomes of the sequence-based CRAC and CARC motifs. Information gleaned from our characterization will eventually enable a structure-based approach for prediction and design of cholesterol binding sites.