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. 2018 Jun 11;7:e35578. doi: 10.7554/eLife.35578

Figure 4. Complementation characterization of EcFtsZ mutants at inter-protofilament lateral interfaces.

Ten EcFtsZ mutants were selected based on our crystallographic observations. Mutations were introduced by altering hydrophilic residues to hydrophobic leucine. The division phenotype was characterized using a ΔftsZ strain expressing the EcFtsZ mutants, with conditional expression of wild-type FtsZ from a plasmid at 30°C but not 42°C. ‘Rep’ and ‘Ind’ indicate repression and induction media, respectively. For each mutant, the complementation assay was repeated three times.

Figure 4.

Figure 4—figure supplement 1. Complementation characterization of pBpa-incorporated EcFtsZ variants.

Figure 4—figure supplement 1.

Ten replacements of EcFtsZ were selected based on our crystallographic observations. pBpa-incorporated variants were created by altering the corresponding sense codon to an amber TAG stop codon. The FtsZ expression plasmid pTet-FtsZ carrying an in-frame amber mutation was used to transform E. coli LY928-ΔftsZ (pJSB100) cells. Cell division, and thus cell growth, could only occur when an FtsZ variant was functional and in the addition of glucose and pBpa.
Figure 4—figure supplement 2. Immunoblotting analysis of three pBpa variants of EcFtsZ that failed to complement the ftsZ conditional-null strain shows absence of crosslinked dimers.

Figure 4—figure supplement 2.

The variants were expressed in LY928-FtsZ-AviTag cells and crosslinked by UV irradiation. Lysate from cells expressing the R78pBpa variant of FtsZ was analyzed as a positive control for photocrosslinked FtsZ dimers.