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Mechanism of Feedback Allosteric Inhibition of ATP Phosphoribosyltransferase

[Image: see text] MtATP-phosphoribosyltransferase catalyzes the first and committed step in l-histidine biosynthesis in Mycobacterium tuberculosis and is therefore subjected to allosteric feedback regulation. Because of its essentiality, this enzyme is being studied as a potential target for novel a...

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Autores principales: Pedreño, Sònia, Pisco, João Pedro, Larrouy-Maumus, Gérald, Kelly, Geoff, de Carvalho, Luiz Pedro Sório
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Chemical Society 2012
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3466779/
https://www.ncbi.nlm.nih.gov/pubmed/22989207
http://dx.doi.org/10.1021/bi300808b
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author Pedreño, Sònia
Pisco, João Pedro
Larrouy-Maumus, Gérald
Kelly, Geoff
de Carvalho, Luiz Pedro Sório
author_facet Pedreño, Sònia
Pisco, João Pedro
Larrouy-Maumus, Gérald
Kelly, Geoff
de Carvalho, Luiz Pedro Sório
author_sort Pedreño, Sònia
collection PubMed
description [Image: see text] MtATP-phosphoribosyltransferase catalyzes the first and committed step in l-histidine biosynthesis in Mycobacterium tuberculosis and is therefore subjected to allosteric feedback regulation. Because of its essentiality, this enzyme is being studied as a potential target for novel anti-infectives. To understand the basis for its regulation, we characterized the allosteric inhibition using gel filtration, steady-state and pre-steady-state kinetics, and the pH dependence of inhibition and binding. Gel filtration experiments indicate that MtATP-phosphoribosyltransferase is a hexamer in solution, in the presence or absence of l-histidine. Steady-state kinetic studies demonstrate that l-histidine inhibition is uncompetitive versus ATP and noncompetitive versus PRPP. At pH values close to neutrality, a K(ii) value of 4 μM was obtained for l-histidine. Pre-steady-state kinetic experiments indicate that chemistry is not rate-limiting for the overall reaction and that l-histidine inhibition is caused by trapping the enzyme in an inactive conformation. The pH dependence of binding, obtained by nuclear magnetic resonance, indicates that l-histidine binds better as the neutral α-amino group. The pH dependence of inhibition (K(ii)), on the contrary, indicates that l-histidine better inhibits MtATP-phosphoribosytransferase with a neutral imidazole and an ionized α-amino group. These results are combined into a model that accounts for the allosteric inhibition of MtATP-phosphoribosyltransferase.
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spelling pubmed-34667792012-10-10 Mechanism of Feedback Allosteric Inhibition of ATP Phosphoribosyltransferase Pedreño, Sònia Pisco, João Pedro Larrouy-Maumus, Gérald Kelly, Geoff de Carvalho, Luiz Pedro Sório Biochemistry [Image: see text] MtATP-phosphoribosyltransferase catalyzes the first and committed step in l-histidine biosynthesis in Mycobacterium tuberculosis and is therefore subjected to allosteric feedback regulation. Because of its essentiality, this enzyme is being studied as a potential target for novel anti-infectives. To understand the basis for its regulation, we characterized the allosteric inhibition using gel filtration, steady-state and pre-steady-state kinetics, and the pH dependence of inhibition and binding. Gel filtration experiments indicate that MtATP-phosphoribosyltransferase is a hexamer in solution, in the presence or absence of l-histidine. Steady-state kinetic studies demonstrate that l-histidine inhibition is uncompetitive versus ATP and noncompetitive versus PRPP. At pH values close to neutrality, a K(ii) value of 4 μM was obtained for l-histidine. Pre-steady-state kinetic experiments indicate that chemistry is not rate-limiting for the overall reaction and that l-histidine inhibition is caused by trapping the enzyme in an inactive conformation. The pH dependence of binding, obtained by nuclear magnetic resonance, indicates that l-histidine binds better as the neutral α-amino group. The pH dependence of inhibition (K(ii)), on the contrary, indicates that l-histidine better inhibits MtATP-phosphoribosytransferase with a neutral imidazole and an ionized α-amino group. These results are combined into a model that accounts for the allosteric inhibition of MtATP-phosphoribosyltransferase. American Chemical Society 2012-09-18 2012-10-09 /pmc/articles/PMC3466779/ /pubmed/22989207 http://dx.doi.org/10.1021/bi300808b Text en Copyright © 2012 American Chemical Society http://pubs.acs.org This is an open-access article distributed under the ACS AuthorChoice Terms & Conditions. Any use of this article, must conform to the terms of that license which are available at http://pubs.acs.org.
spellingShingle Pedreño, Sònia
Pisco, João Pedro
Larrouy-Maumus, Gérald
Kelly, Geoff
de Carvalho, Luiz Pedro Sório
Mechanism of Feedback Allosteric Inhibition of ATP Phosphoribosyltransferase
title Mechanism of Feedback Allosteric Inhibition of ATP Phosphoribosyltransferase
title_full Mechanism of Feedback Allosteric Inhibition of ATP Phosphoribosyltransferase
title_fullStr Mechanism of Feedback Allosteric Inhibition of ATP Phosphoribosyltransferase
title_full_unstemmed Mechanism of Feedback Allosteric Inhibition of ATP Phosphoribosyltransferase
title_short Mechanism of Feedback Allosteric Inhibition of ATP Phosphoribosyltransferase
title_sort mechanism of feedback allosteric inhibition of atp phosphoribosyltransferase
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3466779/
https://www.ncbi.nlm.nih.gov/pubmed/22989207
http://dx.doi.org/10.1021/bi300808b
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