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. 2016 Mar 7;113(12):3335–3340. doi: 10.1073/pnas.1600216113

Fig. 6.

Fig. 6.

Serine dose–response behaviors of prototype interface 2 mutants. In vivo FRET kinase assays were performed in strain UU2784 [∆(cheA-cheZ) ∆(tar, tap, trg, aer)], which lacks the sensory adaptation enzymes (CheR− CheB−) and expresses only the Tsr chemoreceptor. CheA and CheW proteins were expressed from plasmid pPM25 derivatives in combination with the FRET reporter pair (CheY-YFP/CheZ-CFP) from plasmid pVS88. Normalized kinase activities at each serine test concentration were fitted to a multisite Hill equation to obtain K1/2 and Hill coefficient values. The wild type CheA/CheW proteins and the CheA-A546C/CheW-E27C reporter pair produced K1/2 values of 17 µM and 19 µM, respectively with Hill coefficients of 17 and 15, respectively. Corresponding values for interface 2 mutants were as follows: CheW-X3, 8.9 µM and 1.6; CheA-L545S, 6.0 µM and 1.6; CheA-V551A, 9.7 µM and 1.9; CheA-Y558G, 2.7 µM and 1.6.