Molecular Mechanism and Evolution of Guanylate Kinase Regulation by (p)ppGpp

The nucleotide (p)ppGpp mediates bacterial stress responses, but its targets and underlying mechanisms of action vary among bacterial species and remain incompletely understood. Here, we characterize the molecular interaction between (p)ppGpp and guanylate kinase (GMK), revealing the importance of this interaction in adaptation to starvation. Combining structural and kinetic analyses, we show that (p)ppGpp binds the GMK active site and competitively inhibits the enzyme. The (p)ppGpp-GMK interaction prevents the conversion of GMP to GDP, resulting in GMP accumulation upon amino acid downshift. Abolishing this interaction leads to excess (p)ppGpp and defective adaptation to amino acid starvation. A survey of GMKs from phylogenetically diverse bacteria shows that the (p)ppGpp-GMK interaction is conserved in members of Firmicutes, Actinobacteria, and Deinococcus-Thermus, but not in Proteobacteria, where (p)ppGpp regulates RNA polymerase (RNAP). We propose that GMK is an ancestral (p)ppGpp target and RNAP evolved more recently as a direct target in Proteobacteria. [Display omitted] •(p)ppGpp binds the GMK active site and competitively inhibits its activity•GMK regulation by (p)ppGpp is important for adaptation to amino acid starvation•(p)ppGpp-GMK regulation is broadly conserved in bacteria, including many pathogens•The GMK regulation is largely absent in Proteobacteria (p)ppGpp is pivotal to bacterial stress resistance. Liu et al. (2015) present a high-resolution structure of pppGpp-bound GMK, an essential GTP biosynthesis enzyme. They show that (p)ppGpp-GMK regulation is broadly conserved and important for stress adaptation by preventing accumulation of excess (p)ppGpp, providing mechanistic and evolutionary insights into (p)ppGpp-mediated stress resistance.