Fig. 4: KKP is a phosphorylation-based toxin/antitoxin system.
From: Control of lysogeny and antiphage defense by a prophage-encoded kinase-phosphatase module
a Domain organization (top) of KKPMP and toxicity of PAO1 with expression of indicated KKPMP components with or without inducer (bottom). b AlphaFold2 prediction of the heterodimer structure of PfkA-PfkB complex. The identified phosphorylation sites in PfkA and PfKB are labeled in red (Supplementary Fig. 6d). c Heat map showing a subset of proteins with highest fold changes between KKMP versus KKPMP in PAO1 identified by phosphoproteomics (significant hits listed in Supplementary Table 5). d Live-dead staining of planktonic PAO1 cells overexpressing KKMP or KKPMP. Cells were induced with IPTG for 6 h before staining. PI, propidium iodide. e Timelapse microscopy of immobilized E. coli MG1655 cells carrying KKPMP on a two-plasmid expression system, pTac-pfkB-pfkA (IPTG-inducible) and pHERD20T-pfpC (arabinose-inducible). Yellow arrows indicate dividing cells. f Dose-dependence of KKPMP toxicity in E. coli MG1655. pfkA and pfkB were cloned into two-plasmid systems (green and gray circles) with pTac (IPTG-inducible) and pHERD20T (arabinose-inducible) promoters. pTac has a higher leakage expression than that of pHERD20T. The top row without induction showed leaky kinase toxicity. Expression levels were evaluated by qRT-PCR and normalized to 16S rRNA gene expression levels. ND, not detected. Three independent cultures were used in (a, c, d, f), and representative images were shown in (a, d–f), and data are shown as the mean ± SD in (f).
