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https://hdl.handle.net/2440/70932
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Type: | Journal article |
Title: | Probing the allosteric activation of pyruvate carboxylase using 2',3'-O-(2,4,6-trinitrophenyl) adenosine 5'-triphosphate as a fluorescent mimic of the allosteric activator acetyl CoA |
Author: | Adina-Zada, A. Hazra, R. Sereeruk, C. Jitrapakdee, S. Zeczycki, T. Maurice, M. Cleland, W. Wallace, J. Attwood, P. |
Citation: | Archives of Biochemistry and Biophysics, 2011; 509(2):117-126 |
Publisher: | Academic Press Inc |
Issue Date: | 2011 |
ISSN: | 0003-9861 1096-0384 |
Statement of Responsibility: | Abdussalam Adina-Zada, Rasmani Hazra, Chutima Sereeruk, Sarawut Jitrapakdee, Tonya N. Zeczycki, Martin St. Maurice, W. Wallace Cleland, John C. Wallace, Paul V. Attwood |
Abstract: | 2',3'-O-(2,4,6-Trinitrophenyl) adenosine 5'-triphosphate (TNP-ATP) is a fluorescent analogue of ATP. MgTNP-ATP was found to be an allosteric activator of pyruvate carboxylase that exhibits competition with acetyl CoA in activating the enzyme. There is no evidence that MgTNP-ATP binds to the MgATP substrate binding site of the enzyme. At concentrations above saturating, MgATP activates bicarbonate-dependent ATP cleavage, but inhibits the overall reaction. The fluorescence of MgTNP-ATP increases by about 2.5-fold upon binding to the enzyme and decreases on addition of saturating acetyl CoA. However, not all the MgTNP-ATP is displaced by acetyl CoA, or with a combination of saturating concentrations of MgATP and acetyl CoA. The kinetics of the binding of MgTNP-ATP to pyruvate carboxylase have been measured and shown to be triphasic, with the two fastest phases having pseudo first-order rate constants that are dependent on the concentration of MgTNP-ATP. The kinetics of displacement from the enzyme by acetyl CoA have been measured and also shown to be triphasic. A model of the binding process is proposed that links the kinetics of MgTNP-ATP binding to the allosteric activation of the enzyme. |
Keywords: | Pyruvate carboxylase Allosteric activation 20,30-O-(2,4,6-Trinitrophenyl) adenosine 50-triphosphate Fluorescence stopped-flow |
Rights: | Copyright © 2011 Elsevier Inc. All rights reserved. |
DOI: | 10.1016/j.abb.2011.03.006 |
Grant ID: | http://purl.org/au-research/grants/arc/DP0988153 http://purl.org/au-research/grants/arc/DP0988153 |
Published version: | http://dx.doi.org/10.1016/j.abb.2011.03.006 |
Appears in Collections: | Aurora harvest Molecular and Biomedical Science publications |
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