Bacterial proteins that are damaged beyond repair can be targeted for proteolytic destruction by conformational changes that expose amino acid motifs called degrons, by the covalent attachment of polypeptide tags or by the binding of adaptor proteins that are recognized by proteases. In the ClpCP protease complex, ClpC unfolds substrates and feeds them into the active site of the protease ClpP. The adaptor protein MecA mediates the binding of substrates to ClpC. Trentini et al. found that arginine phosphorylation also targets bacterial proteins for degradation by ClpCP in a MecA-independent manner. When Bacillus subtilis cells that expressed an inactive form of the protease complex ClpCP were heat-shocked, many of the substrates that were trapped inside the barrel of the protease complex were phosphorylated on arginine residues. In vitro, ClpCP-mediated degradation of unfolded β-casein required either activity of the arginine kinase McsB or the presence of the adaptor MecA. Free phosphoarginine, but not free phosphate or nonphosphorylated arginine, reduced ClpCP-mediated degradation of β-casein in the presence or absence of MecA, suggesting that phosphoarginine competed with MecA for binding to ClpCP. Indeed, crystallographic analysis showed that free phosphorylated arginine bound to ClpC at a site that overlapped with the binding site for MecA, implying that substrates could be targeted to ClpCP either by phosphoarginine-mediated or by adaptor-mediated binding to ClpC. Residues in ClpC that were critical for phosphoarginine binding were conserved across several other Gram-positive species, including pathogens, and mutating these residues reduced the ability of B. subtilis to recover from heat shock. How heat shock stimulates arginine phosphorylation was not determined, but these findings identify arginine phosphorylation as a general mechanism for targeting proteins for degradation in B. subtilis and suggest that this process could be a potential target for the development of new antibacterial drugs (see Tripathi and Gottesman).
D. B. Trentini, M. J. Suskiewicz, A. Heuck, R. Kurzbauer, L. Deszcz, K. Mechtler, T. Clausen, Arginine phosphorylation marks proteins for degradation by a Clp protease. Nature 539, 48–53 (2016). [PubMed]
A. Tripathi, S. Gottesman, Phosphate on, rubbish out. Nature 539, 38–39 (2016). [Online Journal]