FIG. 3.

In vivo transcriptional activity of mutant M6 mga alleles. A) GusA activity of whole-cell lysates. Production of β-glucuronidase activity was determined for lysates from an mga-deleted Pemm-gusA reporter strain KSM148.174 or B) an mga-deleted Pmga-gusA reporter strain KSM231.310 containing plasmids expressing the following M6 Mga alleles from a constitutive Pspac promoter: wild-type M6 Mga, vector only, the M6 Mga mutants Q10R, Q11A, Q11R, W12A, W12R, R13A, E14A, and L15A. An arbitrary mutant K32A/D33A (no defect) and a known DNA-binding mutant in HTH-4 (defective) are included as controls. GusA units represent a measure of absorbance (A₄₂₀)/protein concentration (μg/ml) and are the average of three independent experiments. C) Western analysis was performed on whole-cell lysates from the above samples using both an anti-Mga antibody for detection of protein levels (top) and an anti-Hsp60 antibody as a control for loading (bottom). D) Activity of mutants expressed from the native promoter. Activity levels for both wild-type and the M6 Mga mutants Q11A, Q11R, and R13A were examined for each allele when expressed from the native mga promoter using the Δmga Pemm-gusA reporter strain (top). Protein production was also determined via Western analysis (bottom) as described above. Percentages indicate level of GusA activity compared to that of the wild-type control.