FIG 5.

K^s associated with overexpression of ycgO and its modulation by K⁺ uptake systems and levels of YcgO in the parent strain and ΔptsN mutant. Shown are the growth rates (μ) of pairs of isogenic strains bearing single-copy integrations of the vector (open circles) and ycgO under the expression control of the P_trc promoter (solid circles) in attB with respect to [K⁺]_e. The predominant K⁺ uptake systems present in the pairs of strains are indicated (A to D). Growth rates of all strains were determined following their growth in K₁, K₂₀, K₄₀, and K₁₁₅ media containing 0.1 mM IPTG. The strain pairs employed are represented by open and solid symbols, respectively: JD726 and JD727 (A), GJ14949 and GJ14950 (B), JD728 and JD729 (C), and JD734 and JD735 (D). (E) Expression levels of a 3× FLAG-tagged YcgO encoded by ycgO_FL in JD723 (ycgO_FL) and JD724 (ΔptsN ycgO_FL). The lane labeled “Nil” contains a whole-cell extract of strain JD624. Cultures of JD624, JD723, and JD724 obtained after growth in K₁ medium to mid-exponential phase were centrifuged and suspended in SDS sample buffer at an A₆₀₀ of 0.002/μl. Equal volumes were loaded onto a 12% SDS-PAGE gel that was processed and immunoblotted with anti-FLAG M2 monoclonal antibody. A nonspecific anti-FLAG immunoreactive material indicative of equal loading is marked with an asterisk.