title: Modulation of nucleotide specificity of thermophilic F(o)F(1)-ATP Synthase by epsilon-subunit.
abstract: The C-terminal two α-helices of the ε-subunit of thermophilic Bacillus F(o)F(1)-ATP synthase (TF(o)F(1)) adopt two conformations: an extended long arm ("up-state") and a retracted hairpin ("down-state"). As ATP becomes poor, ε changes the conformation from the down-state to the up-state and suppresses further ATP hydrolysis. Using TF(o)F(1) expressed in Escherichia coli, we compared TF(o)F(1) with up- and down-state ε in the NTP (ATP, GTP, UTP, and CTP) synthesis reactions. TF(o)F(1) with the up-state ε was achieved by inclusion of hexokinase in the assay and TF(o)F(1) with the down-state ε was represented by εΔc-TF(o)F(1), in which ε lacks C-terminal helices and hence cannot adopt the up-state under any conditions. The results indicate that TF(o)F(1) with the down-state ε synthesizes GTP at the same rate of ATP, whereas TF(o)F(1) with the up-state ε synthesizes GTP at a half-rate. Though rates are slow, TF(o)F(1) with the down-state ε even catalyzes UTP and CTP synthesis. Authentic TF(o)F(1) from Bacillus cells also synthesizes ATP and GTP at the same rate in the presence of adenosine 5'-(β,γ-imino)triphosphate (AMP-PNP), an ATP analogue that has been known to stabilize the down-state. NTP hydrolysis and NTP-driven proton pumping activity of εΔc-TF(o)F(1) suggests similar modulation of nucleotide specificity in NTP hydrolysis. Thus, depending on its conformation, ε-subunit modulates substrate specificity of TF(o)F(1).
authors: ATP Synthesis Regulation Project, International Research Project (ICORP), Japan Science and Technology Corporation, Aomi 2-41, Tokyo 135-0064, Japan.
authors: Toshiharu
authors: T
authors: Suzuki
authors: Chiaki
authors: C
authors: Wakabayashi
authors: Kazumi
authors: K
authors: Tanaka
authors: Boris A
authors: BA
authors: Feniouk
authors: Masasuke
authors: M
authors: Yoshida
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry; D001407:Bacillus / Q000201:enzymology; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004926:Escherichia coli / Q000201:enzymology; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D006160:Guanosine Triphosphate / Q000737:chemistry; D006868:Hydrolysis; D009711:Nucleotides / Q000737:chemistry* / Q000235:genetics; D011487:Protein Conformation; D021122:Protein Subunits / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D006180:Proton-Translocating ATPases / Q000737:chemistry*; D013379:Substrate Specificity; D013696:Temperature; D000097795:ATPase Inhibitory Protein
chemical_list: D000255:Adenosine Triphosphate; D006160:Guanosine Triphosphate; D009711:Nucleotides; D021122:Protein Subunits; D011506:Proteins; D006180:Proton-Translocating ATPases; D000097795:ATPase Inhibitory Protein