Mutational analysis of Era, an essential GTP-binding protein of Escherichia coli.
Era is an essential GTP-binding protein of an unknown function in Escherichia coli. On the basis of its sequence similarities to other GTP-binding proteins such as E. coli EF-Tu, EF-G, IF2 and eukaryotic Ras proteins, it has been suggested that the Era function is activated by GTP binding, and that subsequent conversion of bound GTP to GDP by the intrinsic GTPase activity modulates its function. Two Era mutants, one dominant negative mutant (dE), which has a deletion mutation from Ala40 to Gly49, and the other non-functional mutant (T42A/T43A), which has two substitution mutations, Thr42 to Ala and Thr43 to Ala, were analyzed for their abilities of GTP-binding and GTPase activity. It was found that the dE mutant lost the GTP-binding ability, while it still retained the GTPase activity. On the other hand, the T42A/T43A mutant retained both the GTP-crosslinking and GTPase activities. However, the Km values for GTPase activity increased 5- and 12-fold for dE and T42A/T43A mutants, respectively. These results indicate that both the GTP-binding and GTPase activities are important for the Era function.
Shimamoto
T
T
Department of Biochemistry, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway 08854-5635, USA.
Inouye
M
M
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D003432:Cross-Linking Reagents; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins; D020558:GTP Phosphohydrolases / Q000235:genetics* / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005798:Genes, Bacterial; D006160:Guanosine Triphosphate / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D010766:Phosphorylation; D017354:Point Mutation; D016601:RNA-Binding Proteins; D017384:Sequence Deletion
D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D029968:Escherichia coli Proteins; D016601:RNA-Binding Proteins; C068027:era protein, E coli; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins