Phosphorylation of the calcium adenosine triphosphatase of sarcoplasmic reticulum: rate-limiting conformational change followed by rapid phosphoryl transfer

Petithory, Joanne R.; Jencks, William P.

doi:10.1021/bi00364a006

Cited by 75 publications

(112 citation statements)

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“…This finding is in line with models where the ␥-phosphate in the nucleotide-ATPase complex is still some distance away from the phosphorylation site, as proposed by Hua et al (46) for the Ca 2ϩ -ATPase and Ettrich et al (47) for the Na ϩ /K ϩ -ATPase. Then, the ␥-phosphate arrives at the catalytic site only after nucleotide binding, which could take place during the conformational change after nucleotide binding that has been identified as the rate-limiting step for phosphorylation (48). A ␥-phosphate in some distance to the phosphorylation site provides a possibility of binding a regulatory ATP molecule to the phosphoenzyme at the same site (49).…”

Section: Discussionmentioning

confidence: 99%

Mapping Interactions between the Ca2+-ATPase and Its Substrate ATP with Infrared Spectroscopy

Liu

Barth

2003

Journal of Biological Chemistry

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Infrared spectroscopy has been used to map substrate-protein interactions: the conformational changes of the sarcoplasmic reticulum Ca 2؉ -ATPase upon nucleotide binding and ATPase phosphorylation were monitored using the substrate ATP and ATP analogues (2 -deoxy-ATP, 3 -deoxy-ATP, and inosine 5 -triphosphate), which were modified at specific functional groups of the substrate. Modifications to the 2 -OH, the 3 -OH, and the amino group of adenine reduce the extent of bindinginduced conformational change of the ATPase, with particularly strong effects observed for the latter two. This demonstrates the structural sensitivity of the nucleotide-ATPase complex to individual interactions between nucleotide and ATPase. All groups studied are important for binding and interactions of a given ligand group with the ATPase depend on interactions of other ligand groups.Phosphorylation of the ATPase was observed for ITP and 2 -deoxy-ATP, but not for 3 -deoxy-ATP. There is no direct link between the extent of conformational change upon nucleotide binding and the rate of phosphorylation showing that the full extent of the ATP-induced conformational change is not mandatory for phosphorylation. As observed for the nucleotide-ATPase complex, the conformation of the first phosphorylated ATPase intermediate E1PCa 2 also depends on the nucleotide, indicating that ATPase states have a less uniform conformation than previously anticipated.Ligand binding to proteins controls vast numbers of cellular processes and has attracted great scientific and economic interest. Protein and ligand flexibility are important determinants of the interaction and often lead to ligand binding modes that are not anticipated from structures obtained with other ligands. To these "failure(s) of the rigid receptor hypothesis" (1) is added here an impressive example: induced-fit binding of nucleotides to the Ca 2ϩ -ATPase. This finding stems from a systematic mapping of substrate-protein interactions with infrared (IR) 1 spectroscopy. New approaches like this are welcome in the field of ligand-protein recognition, since the most informative techniques, NMR and x-ray crystallography, are laborious and problematic for some systems. Methods like fluorescence and luminescence that require less expenditure also provide less molecular information. We expect that this technology gap will be bridged by IR spectroscopy.IR spectroscopy, one of the methods of vibrational spectroscopy, provides direct information on the molecular level, is cost-effective, and can be universally applied from small soluble proteins to large membrane proteins under near-physiological conditions. Work summarized in recent reviews (2-5) has shown that the vibrational spectrum changes characteristically when a ligand binds to a protein. This provides a direct observation of ligand binding: no marker compound has to be introduced to report the binding process, as with many other methods. Previous work has mostly focused on individual interactions between a ligand and a protein by monitoring the ...

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Section: Discussionmentioning

confidence: 99%

Mapping Interactions between the Ca2+-ATPase and Its Substrate ATP with Infrared Spectroscopy

Liu

Barth

2003

Journal of Biological Chemistry

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“…Mg . ATP promotes fast phosphorylation when calcium is bound to the transport sites (Froehlich and Taylor, 1975;Verjovski-Almeida and Inesi, 1979;Jencks, 1984,1987;Petithory and Jencks, 1986;Fernandez-Belda and Inesi, 1986), and, at room temperature in the presence of potassium, the reaction steps leading to phosphoenzyme formation are so fast that Mg . ATP binding has not been directly measured.…”

Section: The Reaction Mechanism Ofmentioning

confidence: 99%

“…ATP binding has not been directly measured. Equilibrium and rate constants for ATP binding have been deduced from kinetics of phosphorylation and dephosphorylation (Sumida et al, 1980;Pickart and Jencks, 1984;Fernandez-Belda et al, 1984, 1986Teruel et al, 1987;Stahl and Jencks, 1987;Petithory and Jencks, 1986). However, working at low temperature and in the absence of potassium, we have recently completed a detailed study of the reaction mechanism with Ca .…”

Section: The Reaction Mechanism Ofmentioning

confidence: 99%

The reaction mechanism of Ca²⁺‐ATPase of sarcoplasmic reticulum

Lacapère

Guillain

1993

European Journal of Biochemistry

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Combining rapid filtration and rapid acid quenchmg, we have directly measured, at pH 7.0 and 5 "C, the association and dissociation rate constants of Mg . ATP binding to the sarcoplasmic reticulum (SR) ATPase in the presence of 50 pM calcium and 5 mM MgC12 (3-4x lo6 M-' . s-' and 9 s-', respectively). Therefore, we have determined the true affinity for Mg . ATP (Kd = 3 pM) in the presence of calcium, which can not be measured at equilibrium because of spontaneous and fast phosphorylation. At low concentrations, Mg . ATP binding is the rate limiting step in the phosphorylation process, and Mg . ATP dissociation is slower than dephosphorylation.The kinetics of Ca2+ binding measured by rapid filtration are biphasic, reflecting a two-step mechanism, both steps being accelerated by Mg . ATP. Combining rapid filtration and rapid monitoring of the intrinsic fluorescence of SR Ca2+-ATPase, we showed that rate constants for calcium binding are always lower than those of Mg . ATP binding to an EGTA-incubated enzyme. We measured dissociation and association rate constants of Mg . ATP binding in the absence of calcium (k-1 = 25 s-' and kl = 7.5 lo6 M-' . s-' ). This gives a Kd similar to that obtained by equilibrium measurements (3 -4 pM).Both non-phosphorylated conformations of the enzyme have similar affinity for Mg . ATP.Therefore, activation of ATPase activity by an excess of ATP cannot be explained by a change in affinity of the non-phophorylated enzyme for Mg . ATP. In conjunction with previous results, these data are used to discuss the molecular mechanism for the Ca2+-ATPase cycle, in which ATP is sequentially substrate and activator on a multiple-function single site.Calcium transport by Ca2 +-ATPase of skeletal-muscle sarcoplasmic reticulum (SR) proceeds at the expense of ATP hydrolysis. The reaction mechanism is commonly represented by a scheme in which the enzyme resides in two main conformations: El in the presence of calcium, and E2 in the absence of calcium (reviews: de Meis, 1981;Inesi, 1985).It has already been observed that ATP concentrations higher than that required for phosphoenzyme formation accelerate ATP hydrolysis (Weber et al., 1966). Several hypotheses have been proposed to explain t h s activation process. One proposes that, similar to Na/K-ATPase (for a review, see Skou, 1990) the affinity of the enzyme for Mg . ATP could change depending on the enzyme conformation (Reynolds et al., 1985). Two distinct sites have also been postulated, one catalytic and one regulatory (de Meis and de Mello, 1973;Coll and Murphy, 1985,1991 Abbreviations. SR, sarcoplasmic reticulum; kobsr observed rate constant; k,,, association rate constant; koff, dissociation rate constant; Kp, equilibrium constant of phosphorylation; Vo, initial rate; Np,ATP, 2'(3')-0-(2,4,6-trinitrophenyl)adenosine 5'-triphosphate.Enzyme. Ca-transporting ATPase (EC 3.6

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“…However, it has been suggested previously that a similar occlusion occurs neither after the mere formation of a noncovalent E1⅐AMPPCP complex, nor after the mere formation of the E1⅐Mg⅐ATP complex that immediately precedes phosphorylation during the normal cycle (4,9,12,13). This apparent discrepancy between the different properties of the two ATPase forms and their similar crystalline structure has already been noted (4,5,9), and various interpretations have been given, together with somewhat contradictory comments that these two forms, E1⅐AMPPCP and E1⅐AlFx⅐ADP, have (5) or do not have (4) a similar pattern of resistance to cleavage by proteinase K of their cytosolic domains.…”

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confidence: 95%

The Average Conformation at Micromolar [Ca2+] of Ca2+-ATPase with Bound Nucleotide Differs from That Adopted with the Transition State Analog ADP·AlFx or with AMPPCP under Crystallization Conditions at Millimolar [Ca2+]

Picard

Toyoshima²,

Champeil³

2005

Journal of Biological Chemistry

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Crystalline forms of detergent-solubilized sarcoplasmic reticulum Ca 2؉ -ATPase, obtained in the presence of either a substrate analog, AMPPCP, or a transition state complex, ADP⅐fluoroaluminate, were recently described to share the same general architecture despite the fact that, when studied in a test tube, these forms show different functional properties. Here, we show that the differences in the properties of the E1⅐AMPPCP and the E1⅐ADP⅐AlFx membraneous (or solubilized) forms are much less pronounced when these properties are examined in the presence of 10 mM Ca 2؉ (the concentration prevailing in the crystallization media) than when they are examined in the presence of the few micromolar of Ca 2؉ known to be sufficient to saturate the transport sites. This concerns various properties, including ATPase susceptibility to proteolytic cleavage by proteinase K, ATPase reactivity toward SH-directed Ellman's reagent, ATPase intrinsic fluorescence properties (here described for the E1⅐ADP⅐AlFx complex for the first time), and also the rates of 45 Ca 2؉ -40 Ca 2؉ exchange at site "II." These results solve the above paradox at least partially and suggest that the presence of a previously unrecognized Ca 2؉ ion in the E1⅐AMPPCP crystals should be re-investigated. A contrario, they emphasize the fact that the average conformation of the E1⅐AMPPCP complex under usual conditions in the test tube differs from that found in the crystalline form. The extended conformation of nucleotide revealed by the E1⅐AMPPCP crystalline form might be only indicative of the requirements for further processing of the complex, toward the transition state leading to phosphorylation and Ca 2؉ occlusion.

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Phosphorylation of the calcium adenosine triphosphatase of sarcoplasmic reticulum: rate-limiting conformational change followed by rapid phosphoryl transfer

Cited by 75 publications

References 24 publications

Mapping Interactions between the Ca2+-ATPase and Its Substrate ATP with Infrared Spectroscopy

Mapping Interactions between the Ca2+-ATPase and Its Substrate ATP with Infrared Spectroscopy

The reaction mechanism of Ca²⁺‐ATPase of sarcoplasmic reticulum

The Average Conformation at Micromolar [Ca2+] of Ca2+-ATPase with Bound Nucleotide Differs from That Adopted with the Transition State Analog ADP·AlFx or with AMPPCP under Crystallization Conditions at Millimolar [Ca2+]

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Phosphorylation of the calcium adenosine triphosphatase of sarcoplasmic reticulum: rate-limiting conformational change followed by rapid phosphoryl transfer

Cited by 75 publications

References 24 publications

Mapping Interactions between the Ca2+-ATPase and Its Substrate ATP with Infrared Spectroscopy

Mapping Interactions between the Ca2+-ATPase and Its Substrate ATP with Infrared Spectroscopy

The reaction mechanism of Ca2+‐ATPase of sarcoplasmic reticulum

The Average Conformation at Micromolar [Ca2+] of Ca2+-ATPase with Bound Nucleotide Differs from That Adopted with the Transition State Analog ADP·AlFx or with AMPPCP under Crystallization Conditions at Millimolar [Ca2+]

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The reaction mechanism of Ca²⁺‐ATPase of sarcoplasmic reticulum