Ser/Thr/Tyr protein phosphorylation in the archaeon Halobacterium salinarum--a representative of the third domain of life

Abstract

In the quest for the origin and evolution of protein phosphorylation, the major regulatory post-translational modification in eukaryotes, the members of archaea, the "third domain of life", play a protagonistic role. A plethora of studies have demonstrated that archaeal proteins are subject to post-translational modification by covalent phosphorylation, but little is known concerning the identities of the proteins affected, the impact on their functionality, the physiological roles of archaeal protein phosphorylation/dephosphorylation, and the protein kinases/phosphatases involved. These limited studies led to the initial hypothesis that archaea, similarly to other prokaryotes, use mainly histidine/aspartate phosphorylation, in their two-component systems representing a paradigm of prokaryotic signal transduction, while eukaryotes mostly use Ser/Thr/Tyr phosphorylation for creating highly sophisticated regulatory networks. In antithesis to the above hypothesis, several studies showed that Ser/Thr/Tyr phosphorylation is also common in the bacterial cell, and here we present the first genome-wide phosphoproteomic analysis of the model organism of archaea, Halobacterium salinarum, proving the existence/conservation of Ser/Thr/Tyr phosphorylation in the "third domain" of life, allowing a better understanding of the origin and evolution of the so-called "Nature's premier" mechanism for regulating the functional properties of proteins.

Figure 1. Statistical analysis of the results of the phosphoproteome analysis of H. salinarum.

A. The number of phosphoproteins, phosphoserines, phosphothreonines and phosphotyrosines identified in Wt H. salinarum strain R1, ΔserB, and overall. B. The function classes distribution of the phosphoproteins identified in H. salinarum.

Figure 2. MS/MS spectrum of the phosphopeptide AQDRApTEGEQTAETAIDR of the cytoplasmic arginine transducer protein Car (OE5243F).

The phosphorylation site (Thr245) is located on the methyl-accepting chemotaxis protein (MCP) signal domain of the protein. This is the first time that a MCP is reported to be phosphorylated.

Figure 3. Conservation of protein phosphorylation.

A. Overlap of phosphoproteins detected in B. subtilis, E.coli and H. salinarum. B. Average conservation of phosphorylated (dark-gray) and non-phosphorylated (light-gray) serines that occur in the loop regions. C. Conservation of the identified H. salinarum phosphoproteins in comparison to the whole proteome conservation among the three domains of life.