Regulation of Shigella Effector Kinase OspG through Modulation of Its Dynamic Properties

Gram-negative pathogens secrete effector proteins into human cells to modulate normal cellular processes and establish a bacterial replication niche. Shigella and pathogenic Escherichia coli possess homologous effector kinases, OspG and NleH1/2, respectively. Upon translocation, OspG but not NleH binds to ubiquitin and a subset of E2 similar to Ub conjugates, which was shown to activate its kinase activity. Here we show that OspG, having a minimal kinase fold, acquired a novel mechanism of regulation of its activity. Binding of the E2 similar to Ub conjugate to OspG not only stimulates its kinase activity but also increases its optimal temperature for activity to match the human body temperature and stabilizes its labile C-terminal domain. The melting temperature (T-m) of OspG alone is only 31 degrees C, as compared to 41 degrees C to NleH1/2 homologs. In the presence of E2 similar to Ub, the T-m of OspG increases to similar to 42 degrees C, while Ub by itself increases the T-m to 39 degrees C. Moreover, OspG alone displays maximal activity at 26 degrees C, while in the presence of E2 similar to Ub, maximal activity occurs at similar to 42 degrees C. Using NMR and molecular dynamics calculations, we have identified the C-terminal lobe and, in particular, the C-terminal helix, as the key elements responsible for lower thermal stability of OspG as compared to homologous effector kinases. (C) 2018 Elsevier Ltd. All rights reserved.