ABSTRACT
The aconitase protein of Bacillus subtilis was able to bind specifically to sequences resembling the iron response elements (IREs) found in eukaryotic mRNAs. The sequences bound include the rabbit ferritin IRE and IRE-like sequences in the B. subtilis operons that encode the major cytochrome oxidase and an iron uptake system. IRE binding activity was affected by the availability of iron both in vivo and in vitro. In eukaryotic cells, aconitase-like proteins regulate translation and stability of iron metabolism mRNAs in response to iron availability. A mutant strain of B. subtilis that produces an enzymatically inactive aconitase that was still able to bind RNA sporulated 40x more efficiently than did an aconitase null mutant, suggesting that a nonenzymatic activity of aconitase is important for sporulation. The results support the idea that bacterial aconitases, like their eukaryotic homologs, are bifunctional proteins, showing aconitase activity in the presence of iron and RNA binding activity when cells are iron-deprived.
Subject(s)
Aconitate Hydratase/metabolism , Bacillus subtilis/enzymology , Ferritins/metabolism , RNA-Binding Proteins/metabolism , Aconitate Hydratase/genetics , Animals , Bacillus subtilis/genetics , Bacillus subtilis/physiology , Base Sequence , Consensus Sequence , Ferritins/chemistry , Gene Deletion , Kinetics , Molecular Sequence Data , Mutagenesis , Nucleic Acid Conformation , Oligodeoxyribonucleotides/chemistry , Oligoribonucleotides/chemistry , Oligoribonucleotides/metabolism , Operon , Protein Biosynthesis , RNA, Messenger/metabolism , RNA-Binding Proteins/genetics , Rabbits , Spores, Bacterial , Transcription, GeneticABSTRACT
Two recombinases, XerC and XerD, act at the recombination sites psi and cer in plasmids pSC101 and Co1E1 respectively. Recombination at these sites maintains the plasmids in a monomeric state and helps to promote stable plasmid inheritance. The accessory protein PepA acts at both psi and cer to ensure that only intramolecular recombination takes place. An additional accessory protein, ArgR, is required for recombination at cer but not at psi. Here, we demonstrate that the ArcA/ArcB two-component regulatory system of Escherichia coli, which mediates adaptation to anaerobic growth conditions, is required for efficient recombination in vivo at psi. Phosphorylated ArcA binds to psi in vitro and increases the efficiency of recombination at this site. Binding of ArcA to psi may contribute to the formation of a higher order synaptic complex between a pair of psi sites, thus helping to ensure that recombination is intramolecular.
Subject(s)
Bacterial Outer Membrane Proteins/metabolism , Bacterial Proteins/metabolism , DNA Nucleotidyltransferases/metabolism , DNA-Binding Proteins/metabolism , Escherichia coli Proteins , Escherichia coli/genetics , Integrases , Membrane Proteins/metabolism , Plasmids/genetics , Protein Kinases , Recombination, Genetic , Repressor Proteins , Anaerobiosis , Bacterial Outer Membrane Proteins/genetics , Bacterial Outer Membrane Proteins/isolation & purification , Base Sequence , Binding Sites , DNA Nucleotidyltransferases/genetics , DNA-Binding Proteins/genetics , Escherichia coli/enzymology , Escherichia coli/growth & development , Gene Expression Regulation, Bacterial , Genes, Bacterial , Molecular Sequence Data , Mutation , Phosphorylation , Recombinases , Signal TransductionABSTRACT
Xer site-specific recombination at ColE1 cer converts plasmid multimers into monomers, thus ensuring the heritable stability of ColE1. Two related recombinase proteins, XerC and XerD, catalyse the strand exchange reaction at a 30 bp recombination core site. In addition, two accessory proteins, PepA and ArgR, are required for recombination at cer. These two accessory proteins are thought to act at 180 bp of accessory sequences adjacent to the cer recombination core to ensure that recombination only occurs between directly repeated sites on the same molecule. Here, we demonstrate that PepA and ArgR interact directly with cer, forming a complex in which the accessory sequences of two cer sites are interwrapped approximately three times in a right-handed fashion. We present a model for this synaptic complex, and propose that strand exchange can only occur after the formation of this complex.
