title: High affinity nucleotide-binding mutant of the ε subunit of thermophilic F1-ATPase.
abstract: Specific ATP binding to the ε subunit of thermophilic F1-ATPase has been utilized for the biosensors of ATP in vivo. I report here that the ε subunit containing R103A/R115A mutations can bind ATP with a dissociation constant at 52 nM, which is two orders of magnitude higher affinity than the wild type. The mutant retained specificity for ATP; ADP and GTP bound to the mutant with dissociation constants 16 and 53 μM, respectively. Thus, the mutant would be a good platform for various types of nucleotide biosensor with appropriate modifications.
authors.affiliation: Department of Life Science, College of Science, Rikkyo University, Japan; Research Center for Life Science, College of Science, Rikkyo University, Japan. Electronic address: katoyama@rikkyo.ac.jp.
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Kato-Yamada
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry*; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics; D001665:Binding Sites; D004789:Enzyme Activation; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods*; D021122:Protein Subunits; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D021122:Protein Subunits; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases
keywords: ATP binding; ATP synthase; Biosensor; Inhibitor; Protein engineering