Fig 4.
The P118A replacement in GolS does not affect its stability or its DNA affinity. (A) GolS or GolSP118A was expressed from the GolS-dependent PgolTS promoter in the chromosome by the addition of 40 μM AuHCl4 to the growth medium (LB). Overnight cultures were washed and suspended in with fresh LB without AuHCl4 and incubated for 0, 4, or 12 h. Detection of intracellular GolS or GolSP118A was carried out by Western blotting in whole-cell extracts. (B) Electrophoretic gel mobility shift assay analysis using 6 fmol of 32P 3′-end-labeled PCR fragment from the golB promoter region and the purified GolS variant at the indicated concentrations. (−), no protein addition.