HypB specifically binds (p)ppGpp with physiological affinity. (A) Competition assay of HypB (20 μM) binding α- 32 P-labeled ppGpp and pppGpp (2 nM [each]) in the presence of cold competitors (100 μM). (B) Binding curves and K d determination for HypB binding α- 32 P-labeled ppGpp, pppGpp, and GTP (2 nM [each]). Three replicates were performed, and the apparent K d values are indicated.