Meta‐Proteomic Analysis of PTMs in G protein Signaling Systems – New Tools and Unsuspected Discoveries

Abstract

The collective structural and functional data for prototypical G protein signaling (GPS) systems – defined by 7‐transmembrane receptors (GPCRs), heterotrimeric G proteins (Gαβγ) and Regulators of G‐protein Signaling (RGS) – is arguably the most comprehensive of its kind to date. Comparatively little is known about how G protein structure and function is altered by post‐translational modifications (PTMs). Here we report the first structurally‐resolved, meta‐proteomic map of all empirically validated PTMs for GPS proteins. Using a novel PTM analysis tool called Structural Projection of PTMs (SPoP), a total of 1,910 unique eukaryotic PTMs were coalesced onto available structures within the Protein Data Bank. The resulting dataset has enabled the first‐ever visualization of the PTM topology for each member of a prototypical GPS system. By factoring total PTM observations, protein structure, protein interactions and sequence conservation, we estimated a function potential value for each PTM. Rank order comparison of calculated function potentials with literature‐based evidence of function reveals that SPoP has predictive power. Based on these results, we have identified major PTM hotspots in GPS proteins; predicted and tested the involvement of previously unsuspected PTM regulatory elements; and revealed surprising differences between the PTM topologies of large and small guanine‐nucleotide binding proteins. We propose that meta‐proteomic and structural PTM analysis with SPoP can lead to the discovery of functionally important modification sites that regulate biomedically‐relevant classes of signaling molecules.