Reaction of Bacillus subtilis glutamine phosphoribosylpyrophosphate amidotransferase with oxygen: chemistry and regulation by ligands

Abstract

The inactivation of glutamine phosphoribosylpyrophosphate amidotransferase by reaction of its iron-sulfur center with O2 is believed to be a physiologically important mode of regulation of this enzyme in Bacillus subtilis cells in the stationary phase of growth. Chemical and physical changes accompanying oxidation of the purified enzyme by O2 were studied. The iron of the 4Fe-4S center was oxidized to enzyme-bound high-spin Fe3+; the S2- was oxidized to a mixture of S0 bound as thiocystine and unidentified products. The oxidant appeared to be O2, rather than peroxide, superoxide, hydroxyl radical, or singlet oxygen. Gross physical changes in the oxidized enzyme were shown by its aggregation, decreased solubility, and altered circular dichroic spectrum. Experimental variables affecting the rate of oxidative inactivation were described; the most important of these was modulation of rates of inactivation by the allosteric inhibitors AMP, ADP, GMP, GDP and by the substrate P-Rib-PP. AMP was a potent stabilizer, whose effect was antagonized by P-Rib-PP. The other nucleotides, either acting singly or acting as synergistic pairs, were destabilizers and able to antagonize stabilization by AMP. The results are discussed in terms of the regulation of the stability of amidotransferase and its degradation in vivo.

Roles of the three transcriptional attenuators of the Bacillus subtilis pyrimidine biosynthetic operon in the regulation of its expression

“…This speculative model predicts that the shutoff in pyr transcription during early stationary phase is due to an increase in intracellular UMP pools, a decrease in intracellular PRPP pools, or both. A very sharp decrease in the PRPP pool has been shown to occur upon glucose exhaustion of B. subtilis cells grown on a different medium (5). An alternative mechanism for increased repression by the PyrR protein during sporulation could involve posttranslational modification, such as phosphorylation, as has been found in other attenuation systems (10,11,24).…”

Whole-Genome Transcriptional Analysis of Heavy Metal Stresses in Caulobacter crescentus

“…In addition, glutamate synthase contains an iron-sulfur cluster and has close structural homology with glutamine phosphoribosylpyrophosphate amidotransferase (71). This latter enzyme from B. subtilis is known to be inactivated by O 2 in stationary phase (9). Although up-regulation of ntrBC and PII usually indicates nitrogen limitation response, the key enzyme (glutamine synthetase) was not up-regulated.…”

The LIM region of a presumptive Caenorhabditis elegans transcription factor is an iron-sulfur- and zinc-containing metallodomain.

“…For example, the Krebs cycle enzyme aconitase, which catalyzes the interconversion of citrate and isocitrate, is active in the reduced state and inactive in the oxidized state (17). Similarly, glutamine phosphoribosylpyrophosphate amidotransferase, the initial enzyme ofpurine biosynthesis in Bacillus subtilis, is rapidly inactivated upon oxidation (18 (19,20 (24) and paired (25) families. Binding of DNA by the POU proteins involves both the POU-specific region and the POU homeodomain: although the POU domain does not seem to bind DNA alone, deletion of the POU-specific region decreases DNA binding affinity by a factor of 1000 (26,27).…”