ATP Analogs

Yount, Ralpa G.

doi:10.1002/9780470122884.ch1

Cited by 90 publications

(11 citation statements)

References 187 publications

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“…Furthermore, PPi or AMP-PNP needs to act on this presumed, barely visible closed state at an extremely fast rate; otherwise, this closed event will become more discernible in the single-channel recording. As the chemical structure of AMP-PNP is nearly identical to that of ATP (Yount, 1975), and this ATP analogue can be found comfortably nestled in the ATP-binding sites of ABC transporters (Dawson and Locher, 2007), one comes to an ineluctable implication that ATP can also readily reopen the channel from state X. Thus, contrary to the long-held idea that ATP opens CFTR solely through the long interburst closure, one has to conclude that some of the short-lived closures that are buried in an opening burst can respond to ATP to enter the open state.…”

Section: Discussionmentioning

confidence: 99%

Identification of a novel post-hydrolytic state in CFTR gating

Jih

Saito

et al. 2012

Journal of General Physiology

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Adenosine triphosphate (ATP)-binding cassette (ABC) transporters, ubiquitous proteins found in all kingdoms of life, catalyze substrates translocation across biological membranes using the free energy of ATP hydrolysis. Cystic fibrosis transmembrane conductance regulator (CFTR) is a unique member of this superfamily in that it functions as an ATP-gated chloride channel. Despite difference in function, recent studies suggest that the CFTR chloride channel and the exporter members of the ABC protein family may share an evolutionary origin. Although ABC exporters harness the free energy of ATP hydrolysis to fuel a transport cycle, for CFTR, ATP-induced dimerization of its nucleotide-binding domains (NBDs) and subsequent hydrolysis-triggered dimer separation are proposed to be coupled, respectively, to the opening and closing of the gate in its transmembrane domains. In this study, by using nonhydrolyzable ATP analogues, such as pyrophosphate or adenylyl-imidodiphosphate as baits, we captured a short-lived state (state X), which distinguishes itself from the previously identified long-lived C2 closed state by its fast response to these nonhydrolyzable ligands. As state X is caught during the decay phase of channel closing upon washout of the ligand ATP but before the channel sojourns to the C2 closed state, it likely emerges after the bound ATP in the catalysis-competent site has been hydrolyzed and the hydrolytic products have been released. Thus, this newly identified post-hydrolytic state may share a similar conformation of NBDs as the C2 closed state (i.e., a partially separated NBD and a vacated ATP-binding pocket). The significance of this novel state in understanding the structural basis of CFTR gating is discussed.

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Section: Discussionmentioning

confidence: 99%

Identification of a novel post-hydrolytic state in CFTR gating

Jih

Saito

et al. 2012

Journal of General Physiology

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“…Although p[NH]ppA and ATPγS are frequently employed as non‐hydrolysable analogues of ATP, the precise structures of the three nucleotides do differ. For example, p[NH]ppA has distinct bond lengths (P–N = 1.7 Å compared to P–O = 1.6 Å) and bond angles (P–N–P = 127° compared to P–O–P = 130°), as well as a lower p K a for the terminal phosphate compared to ATP [53]. These stereochemical differences could manifest as non‐identical effects of ATP compared to p[NH]ppA, as has been seen in the F1‐ATPase, where a 100‐fold difference in apparent affinity for ATP and p[NH]ppA binding has been described [54,55].…”

Section: Atp Binding Provides the Initial Impetus For Translocationmentioning

confidence: 99%

The translocation mechanism of P‐glycoprotein

2005

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Multidrug transporters are involved in mediating the failure of chemotherapy in treating several serious diseases. The archetypal multidrug transporter P-glycoprotein (P-gp) confers resistance to a large number of chemically and functionally unrelated anti-cancer drugs by mediating efflux from cancer cells. The ability to efflux such a large number of drugs remains a biological enigma and the lack of mechanistic understanding of the translocation pathway used by P-gp prevents rational design of compounds to inhibit its function. The translocation pathway is critically dependent on ATP hydrolysis and drug interaction with P-gp is possible at one of a multitude of allosterically linked binding sites. However, aspects such as coupling stoichiometry, molecular properties of binding sites and the nature of conformational changes remain unresolved or the centre of considerable controversy. The present review attempts to utilise the available data to generate a detailed sequence of events in the translocation pathway for this dexterous protein.

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“…Kinases can use ATPγS to thiophosphorylate proteins, making them resistant to rapid dephosphorylation. ATPγS can be hydrolyzed by some enzymes but usually at a much lower rate compared with ATP (Yount 1975). Addition of ATPγS to in vitro splicing reactions was initially reported to stall splicing during the second catalytic step (Tazi et al 1992).…”

Section: Introductionmentioning

confidence: 99%

ATPγS stalls splicing after B complex formation but prior to spliceosome activation

Agafonov

Santen

Kastner

et al. 2016

RNA

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The ATP analog ATPγS inhibits pre-mRNA splicing in vitro, but there have been conflicting reports as to which step of splicing is inhibited by this small molecule and its inhibitory mechanism remains unclear. Here we have dissected the effect of ATPγS on pre-mRNA splicing in vitro. Addition of ATPγS to splicing extracts depleted of ATP inhibited both catalytic steps of splicing. At ATPγS concentrations ≥0.5 mM, precatalytic B complexes accumulate, demonstrating a block prior to or during the spliceosome activation stage. Affinity purification of the ATPγS-stalled B complexes (B ATPγS ) and subsequent characterization of their abundant protein components by 2D gel electrophoresis revealed that B ATPγS complexes are compositionally more homogeneous than B complexes previously isolated in the presence of ATP. In particular, they contain little or no Prp19/CDC5L complex proteins, indicating that these proteins are recruited after assembly of the precatalytic spliceosome. Under the electron microscope, B ATPγS complexes exhibit a morphology highly similar to B complexes, indicating that the ATPγS-induced block in the transformation of the B to B act complex is not due to a major structural defect. Likely mechanisms whereby ATPγS blocks spliceosome assembly at the activation stage, including inhibition of the RNA helicase Brr2, are discussed. Given their more homogeneous composition, B complexes stalled by ATPγS may prove highly useful for both functional and structural analyses of the precatalytic spliceosome and its conversion into an activated B act spliceosomal complex.

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ATP Analogs

Cited by 90 publications

References 187 publications

Identification of a novel post-hydrolytic state in CFTR gating

Identification of a novel post-hydrolytic state in CFTR gating

The translocation mechanism of P‐glycoprotein

ATPγS stalls splicing after B complex formation but prior to spliceosome activation

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