The Escherichia coli SeqA protein destabilizes mutant DnaA204 protein

In wild-type Escherichia coli cells, initiation of DNA replication is tight ly coupled to cell growth. In slowly growing dnaA204 (Ts) mutant cells, the cell mass at initiation and its variability is increased two- to threefold relative to wild type. Here, we show that the DnaA protein concentration w as two- to threefold lower in the dnaA204 mutant compared with the wildtype strain. The reason for the DnaA protein deficiency was found to be a rapid degradation of the mutant protein. Absence of SeqA protein stabilized the DnaA204 protein, increased the DnaA protein concentration and normalized th e initiation mass in the dnaA204 mutant cells. During rapid growth, the dna A204 mutant displayed cell cycle parameters similar to wild-type cells as w ell as a normal DnaA protein concentration, even though the DnaA204 protein was highly unstable. Apparently, the increased DnaA protein synthesis comp ensated for the protein degradation under these growth conditions, in which the doubling time was of the same order of magnitude as the half-life of t he protein. Our results suggest that the DnaA204 protein has essentially wi ld-type activity at permissive temperature but, as a result of instability, the protein is present at lower concentration under certain growth conditi ons. The basis for the stabilization in the absence of SeqA is not known. W e suggest that the formation of stable DnaA-DNA complexes is enhanced in th e absence of SeqA, thereby protecting the DnaA protein from degradation.