Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies

Dihydroxyacetone (DHA) kinase from Citrobacter freundii provides an easy entry for the preparation of DHA phosphate; a very important C3 building block in nature. To modify the phosphoryl donor specificity of this enzyme from ATP to inorganic polyphosphate (poly-P); a directed evolution program has...

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Main Authors:	Sánchez-Moreno, Israel, Bordes, Isabel, Castillo, Raquel, Ruiz-Pernía, José Javier, Moliner, Vicent, García-Junceda, Eduardo
Format:	Online
Language:	English
Published:	MDPI 2015
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4661931/

id	pubmed-4661931
recordtype	oai_dc
spelling	pubmed-46619312015-12-10 Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies Sánchez-Moreno, Israel Bordes, Isabel Castillo, Raquel Ruiz-Pernía, José Javier Moliner, Vicent García-Junceda, Eduardo Article Dihydroxyacetone (DHA) kinase from Citrobacter freundii provides an easy entry for the preparation of DHA phosphate; a very important C3 building block in nature. To modify the phosphoryl donor specificity of this enzyme from ATP to inorganic polyphosphate (poly-P); a directed evolution program has been initiated. In the first cycle of evolution, the native enzyme was subjected to one round of error-prone PCR (EP-PCR) followed directly (without selection) by a round of DNA shuffling. Although the wild-type DHAK did not show activity with poly-P, after screening, sixteen mutant clones showed an activity with poly-phosphate as phosphoryl donor statistically significant. The most active mutant presented a single mutation (Glu526Lys) located in a flexible loop near of the active center. Interestingly, our theoretical studies, based on molecular dynamics simulations and hybrid Quantum Mechanics/Molecular Mechanics (QM/MM) optimizations, suggest that this mutation has an effect on the binding of the poly-P favoring a more adequate position in the active center for the reaction to take place. MDPI 2015-11-24 /pmc/articles/PMC4661931/ /pubmed/26610480 http://dx.doi.org/10.3390/ijms161126073 Text en © 2015 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons by Attribution (CC-BY) license (http://creativecommons.org/licenses/by/4.0/).
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Sánchez-Moreno, Israel Bordes, Isabel Castillo, Raquel Ruiz-Pernía, José Javier Moliner, Vicent García-Junceda, Eduardo
spellingShingle	Sánchez-Moreno, Israel Bordes, Isabel Castillo, Raquel Ruiz-Pernía, José Javier Moliner, Vicent García-Junceda, Eduardo Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies
author_facet	Sánchez-Moreno, Israel Bordes, Isabel Castillo, Raquel Ruiz-Pernía, José Javier Moliner, Vicent García-Junceda, Eduardo
author_sort	Sánchez-Moreno, Israel
title	Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies
title_short	Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies
title_full	Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies
title_fullStr	Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies
title_full_unstemmed	Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies
title_sort	tuning the phosphoryl donor specificity of dihydroxyacetone kinase from atp to inorganic polyphosphate. an insight from computational studies
description	Dihydroxyacetone (DHA) kinase from Citrobacter freundii provides an easy entry for the preparation of DHA phosphate; a very important C3 building block in nature. To modify the phosphoryl donor specificity of this enzyme from ATP to inorganic polyphosphate (poly-P); a directed evolution program has been initiated. In the first cycle of evolution, the native enzyme was subjected to one round of error-prone PCR (EP-PCR) followed directly (without selection) by a round of DNA shuffling. Although the wild-type DHAK did not show activity with poly-P, after screening, sixteen mutant clones showed an activity with poly-phosphate as phosphoryl donor statistically significant. The most active mutant presented a single mutation (Glu526Lys) located in a flexible loop near of the active center. Interestingly, our theoretical studies, based on molecular dynamics simulations and hybrid Quantum Mechanics/Molecular Mechanics (QM/MM) optimizations, suggest that this mutation has an effect on the binding of the poly-P favoring a more adequate position in the active center for the reaction to take place.
publisher	MDPI
publishDate	2015
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4661931/
_version_	1613506956853510144

Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies

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