FIG 9.

ZipA Q280L is an intragenic suppressor of FtsZ-binding mutants. (A) Cultures of strain WM5337 (zipA1) transformed with pDSW210-GFP (vector), pDSW210-ZipA-GFP, or derived plasmids carrying zipA mutations were serially diluted, spotted on plates containing 0, 10, 25, 100, or 1,000 μM IPTG, and then grown at permissive (30°C) or restrictive (37°, 42°C) temperatures. The Q280L mutation partially restored ZipA function in combination with the V249E, F269S, and Q290L mutations. Panels show composite images of two plates. (B) Cell lysates of strain WM1074 carrying pUltra-pBpF, pKG110-FtsZ, and pDSW210-ZipA-GFP or derived constructs containing indicated mutations were assessed by Western blotting in triplicate. The Q280L mutation did not significantly affect cross-linking at either the canonical (M226X) or noncanonical (R314X) sites. The mean and standard error (SE) of cross-linking efficiency relative to the control are indicated below the +UV lanes. Values were normalized by total protein levels and ZipA-GFP expression for each strain and then averaged across three replicates. Each panel is a composite image of a single representative blot (replicates not shown).