Adenylosuccinate synthase (ASS) from Saccharomyces cerevisiae has been shown to bind specifically to the T‐rich side of the autonomously replicating sequence (ARS) core consensus sequence [Zeidler, R., Hobert, O., Johannes, L., Faulhammer, H. & Krauss, G. (1993) J. Biol. Chem. 268, 20191–20197]. We have cloned and sequenced the gene for ASS and have studied in detail the enzymatic properties and DNA‐binding activity of ASS. The deduced amino acid sequence of the yeast ASS is highly similar to the same enzymes from other sources from which it is however distinguished by its more basic nature. We show that the enzymatic activity of ASS is inhibited in a highly specific manner by the binding of a 44‐base DNA oligonucleotide carrying the ARS core consensus sequence. Other nucleic acids, rNTP and dNTP are not able to mimic the specific inhibitory effect. Single‐base substitutions in the ARS core sequence lead to a tenfold reduction in inhibition. The inhibition data corroborate the earlier report on the DNA‐binding specificity of this enzyme. The homologous enzymes from Escherichia coli and Dictyostelium discoideum do not show specific binding to single‐stranded ARS sequences and their enzymatic activity is not influenced by the presence of a 44‐base DNA oligonucleotide carrying the ARS core consensus sequence. Treatment of ASS with alkaline phosphatase leads to a loss of DNA binding and to a loss of the inhibition by DNA of the enzymatic activity which suggests that the DNA‐binding activity but not the enzymatic activity may be regulated by the phosphorylation status of the protein.

The ftsH gene, present in all eubacterial species, is anchored in the cytoplasmic membrane and contains an ATP- and a Zn-binding domain that are both part of a metalloprotease activity. The Bacillus subtilis ftsH is not essential, but null mutants exhibit a pleiotropic phenotype including filamentous growth; hypersensitivity towards heat and salt stress and a failure to sporulate. To find out whether one or the other functional domain is involved in these different phenotypes, point mutations were introduced into the coding region for both domains leading to a replacement of conserved amino acid residues. The mutant alleles were fused to a xylose-inducible promoter and integrated ectopically into two different strains, one expressing the wild-type ftsH allele and the other carrying a ftsH knockout. While none of the strains exhibited a growth defect in rich medium at 37 degrees C, those strains expressing only the mutant alleles did not resume growth after heat or salt stress challenge. Furthermore, none of the mutant alleles promoted sporulation. While only those purified mutant FtsH proteins with an intact Walker A box exhibited ATPase activity, all of them failed to degrade beta-casein.