title: Effect of nucleotides on the thermal stability and on the deuteration kinetics of the thermophilic F0F1 ATP synthase.
abstract: Differential scanning calorimetry has been used to characterize the influence of specific nucleotide binding on the thermal unfolding of the F0F1-type ATP synthase from the thermophilic Bacillus PS3 (TF0F1). The calorimetric trace shows an irreversible and kinetically controlled endothermic transition for TF0F1 in the absence of nucleotides. The thermal denaturation occurs at a transition temperature (t(m)) of 81.7 degrees C. The remarkable thermostability of this enzyme was decreased upon tight binding of Mg2+ x ATP to noncatalytic sites, whereas binding of Mg2+ x ADP increased the temperature at which thermal denaturation occurred. At high temperatures, an exothermic transition due to aggregation processes was also affected by nucleotide binding. With the aim to correlate these thermal effects with possible structural differences among the various forms of TF0F1, Fourier transform infrared spectroscopy was carried out. Hydrogen/deuterium exchange was clearly affected by specific nucleotide occupancy. As illustrated by the total extent of protons exchanged, our results demonstrate that more peptide groups are exposed to the medium in the presence of Mg2+ x ATP than in the presence of Mg2+ x ADP. Therefore, consistent with microcalorimetric data, binding of Mg2+ x ADP induces conformational changes which shield amide protons to more buried hydrogen-bonded structures, whereas binding of Mg2+ x ATP results in a more open or flexible structure.
authors.affiliation: Departament de Bioquímica i Biologia Molecular, Facultat de Medicina, Universitat Autònoma de Barcelona, Bellaterra, Spain.
authors.forename: J
authors.initials: J
authors.lastname: Villaverde
authors.forename: J
authors.initials: J
authors.lastname: Cladera
authors.forename: E
authors.initials: E
authors.lastname: Padrós
authors.forename: J L
authors.initials: JL
authors.lastname: Rigaud
authors.forename: M
authors.initials: M
authors.lastname: Duñach
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D001407:Bacillus / Q000201:enzymology; D002152:Calorimetry, Differential Scanning; D003902:Detergents / Q000494:pharmacology; D003903:Deuterium; D004795:Enzyme Stability / Q000187:drug effects; D006860:Hydrogen Bonding; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D011487:Protein Conformation / Q000187:drug effects; D011489:Protein Denaturation / Q000187:drug effects; D017433:Protein Structure, Secondary / Q000187:drug effects; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000378:metabolism*; D017550:Spectroscopy, Fourier Transform Infrared; D013696:Temperature; D013816:Thermodynamics
chemical_list: D003902:Detergents; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D003903:Deuterium; D006180:Proton-Translocating ATPases; D008274:Magnesium