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Int. J. Mol. Sci. 2015, 16(11), 27835-27849; doi:10.3390/ijms161126073

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

1
Departamento de Química Bioorgánica, Instituto de Química Orgánica General, CSIC. Juan de la Cierva 3, Madrid 28006, Spain
2
Departament de Química Física i Analítica, Universitat Jaume I. Castellón 12071, Spain
Current address: Venter Pharma S.L., Azalea 1, Alcobendas 28109, Madrid, Spain
*
Authors to whom correspondence should be addressed.
Academic Editor: Vladimír Křen
Received: 2 October 2015 / Revised: 5 November 2015 / Accepted: 9 November 2015 / Published: 24 November 2015
(This article belongs to the Special Issue Molecular Biocatalysis)
View Full-Text   |   Download PDF [4715 KB, uploaded 24 November 2015]   |  

Abstract

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. View Full-Text
Keywords: biocatalysis; computational chemistry; DHAP-dependent aldolases; dihydroxyacetone kinase; enzyme directed evolution; quantum mechanics/molecular mechanics biocatalysis; computational chemistry; DHAP-dependent aldolases; dihydroxyacetone kinase; enzyme directed evolution; quantum mechanics/molecular mechanics
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Sánchez-Moreno, I.; Bordes, I.; Castillo, R.; Ruiz-Pernía, J.J.; Moliner, V.; García-Junceda, E. Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies. Int. J. Mol. Sci. 2015, 16, 27835-27849.

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