Mutagenesis of ligands to the [4 Fe-4S] center of Bacillus subtilis glutamine phosphoribosylpyrophosphate amidotransferase.
Several mutations were constructed in residues thought to provide ligands for a [4Fe-4S] cluster in Bacillus subtilis amidophosphoribosyltransferase using site-directed mutagenesis of cloned purF. These replacements confirm the identification of cysteinyl ligands to the Fe-S center. Of five mutant enzymes, two had no activity, two less than 25% of the wild type activity, and one was lethal and could not be studied. The Fe content of the two mutant enzymes with partial activity was similar to that of the wild type. Results of partial characterization suggest that the [4Fe-4S] cluster is not involved in allosteric regulation and does not play a specific role in the ammonia- or glutamine-dependent reactions of the enzyme. At least partial enzymatic activity is required for NH2-terminal processing. Pulse labeling experiments suggest that processing is a slow post-translational process which is dependent upon cellular factors. A relationship between Fe-S centers and NH2-terminal processing of an undecapeptide leader suggests a functional connection between these two structural elements in amidophosphoribosyltransferase.
D000582:Amidophosphoribosyltransferase / Q000378:metabolism*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D003553:Cystine / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000378:metabolism*; D008024:Ligands / Q000378:metabolism; D008667:Metalloproteins / Q000378:metabolism*; D009154:Mutation; D010430:Pentosyltransferases / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013329:Structure-Activity Relationship
D000596:Amino Acids; D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D008024:Ligands; D008667:Metalloproteins; D012694:Serine; D003553:Cystine; D010430:Pentosyltransferases; D000582:Amidophosphoribosyltransferase