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F1F0 ATP Synthase (f1f0 + atp_synthase)

Distribution by Scientific Domains

Chemistry	75%

Selected Abstracts

Focused proteomics: Monoclonal antibody-based isolation of the oxidative phosphorylation machinery and detection of phosphoproteins using a fluorescent phosphoprotein gel stain

ELECTROPHORESIS, Issue 15 2004
James Murray
Abstract We have raised monoclonal antibodies capable of immunocapturing all five complexes involved in oxidative phosphorylation for evaluating their post-translational modifications. Complex I (NADH dehydrogenase), complex II (succinate dehydrogenase), complex III (cytochrome c reductase), complex IV (cytochrome c oxidase), and complex V (F1F0 ATP synthase) from bovine heart mitochondria were obtained in good yield from small amounts of tissue in more than 90% purity in one step. The composition and purity of the complexes was evaluated by Western blotting using monoclonal antibodies against individual subunits of the five complexes. In this first study, the phosphorylation state of the proteins without inducing phosphorylation or dephosphorylation was identified by using the novel Pro-Q Diamond phosphoprotein gel stain. The major phosphorylated components were the same as described before in sucrose gradient enriched complexes. In addition a few additional potential phosphoproteins were observed. Since the described monoclonal antibodies show cross reactivity to human proteins, this procedure will be a fast and efficient way of studying post-translational modifications in control and patient samples using only small amounts of tissue. [source]

Membrane embedded location of Na+ or H+ binding sites on the rotor ring of F1F0 ATP synthases

FEBS JOURNAL, Issue 22 2002
Christoph Von Ballmoos
Recent crosslinking studies indicated the localization of the coupling ion binding site in the Na+ -translocating F1F0 ATP synthase of Ilyobacter tartaricus within the hydrophobic part of the bilayer. Similarly, a membrane embedded H+ -binding site is accepted for the H+ -translocating F1F0 ATP synthase of Escherichia coli. For a more definite analysis, we performed parallax analysis of fluorescence quenching with ATP synthases from both I. tartaricus and E. coli. Both ATP synthases were specifically labelled at their c subunit sites with N -cyclohexyl- N, -(1-pyrenyl)carbodiimide, a fluorescent analogue of dicyclohexylcarbodiimide and the enzymes were reconstituted into proteoliposomes. Using either soluble quenchers or spinlabelled phospholipids, we observed a deeply membrane embedded binding site, which was quantitatively determined for I. tartaricus and E. coli to be 1.3 ± 2.4 Å and 1.8 ± 2.8 Å from the bilayer center apart, respectively. These data show a conserved topology among enzymes of different species. We further demonstrated the direct accessibility for Na+ ions to the binding sites in the reconstituted I. tartaricus c11 oligomer in the absence of any other subunits, pointing to intrinsic rotor channels. The common membrane embedded location of the binding site of ATP synthases suggest a common mechanism for ion transfer across the membrane. [source]

Structure of the nucleotide-binding subunit B of the energy producer A1A0 ATP synthase in complex with adenosine diphosphate

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2008
Anil Kumar
A1A0 ATP synthases are the major energy producers in archaea. Like the related prokaryotic and eukaryotic F1F0 ATP synthases, they are responsible for most of the synthesis of adenosine triphosphate. The catalytic events of A1A0 ATP synthases take place inside the A3B3 hexamer of the A1 domain. Recently, the crystallographic structure of the nucleotide-free subunit B of Methanosarcina mazei Gö1 A1A0 ATP synthase has been determined at 1.5,Å resolution. To understand more about the nucleotide-binding mechanism, a protocol has been developed to crystallize the subunit B,ADP complex. The crystallographic structure of this complex has been solved at 2.7,Å resolution. The ADP occupies a position between the essential phosphate-binding loop and amino-acid residue Phe149, which are involved in the binding of the antibiotic efrapeptin in the related F1F0 ATP synthases. This trapped ADP location is about 13,Å distant from its final binding site and is therefore called the transition ADP-binding position. In the trapped ADP position the structure of subunit B adopts a different conformation, mainly in its C-terminal domain and also in the final nucleotide-binding site of the central ,,-domain. This atomic model provides insight into how the substrate enters into the nucleotide-binding protein and thereby into the catalytic A3B3 domain. [source]