Identification of a Novel Class of Farnesylation Targets by Structure-Based Modeling of Binding Specificity

Abstract

Farnesylation is an important post-translational modification catalyzed by farnesyltransferase (FTase). Until recently it was believed that a C-terminal CaaX motif is required for farnesylation, but recent experiments have revealed larger substrate diversity. In this study, we propose a general structural modeling scheme to account for peptide binding specificity and recapitulate the experimentally derived selectivity profile of FTase in vitro. In addition to highly accurate recovery of known FTase targets, we also identify a range of novel potential targets in the human genome, including a new substrate class with an acidic C-terminal residue (CxxD/E). In vitro experiments verified farnesylation of 26/29 tested peptides, including both novel human targets, as well as peptides predicted to tightly bind FTase. This study extends the putative range of biological farnesylation substrates. Moreover, it suggests that the ability of a peptide to bind FTase is a main determinant for the farnesylation reaction. Finally, simple adaptation of our approach can contribute to more accurate and complete elucidation of peptide-mediated interactions and modifications in the cell.

Author Summary

Linear sequence motifs serve as recognition sites for protein-protein interactions as well as for post-translational modifications. One such motif is the CaaX box located at protein C-termini that serves as prenylation site. This prenylation is critical for many signal transduction related proteins and it is thus an important goal to uncover the range of prenylated proteins. Due to poor generalization ability, sequence based computational methods can only go so far in predicting novel targets. In this study, we introduce a novel structure based modeling approach that allows both recovery of known farnesylation substrates, as well as detection of a new class of farnesylation targets. We demonstrate high accuracy in retrospective discrimination between substrates and non-substrates of farnesyltransferase (FTase). More importantly, in a prospective study, in vitro experiments validate that 26/29 predicted peptides indeed undergo farnesylation. These novel peptides were derived either from actual human proteins, or predicted to bind particularly well to FTase. Other than the discovery of putative novel farnesylation targets in the human genome, as well as possible inhibitors, we provide insights into the main determinants of farnesylation. Our approach could be easily extended to additional peptide-protein interactions and help the elucidation of the cellular peptide-protein interaction network.