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Pyranose dehydrogenase ligand promiscuity: a generalized approach to simulate monosaccharide solvation, binding, and product formation.

Graf MM, Zhixiong L, Bren U, Haltrich D, van Gunsteren WF, Oostenbrink C - PLoS Comput. Biol. (2014)

Bottom Line: The free energy difference between β- and α-anomers (ΔGβ-α) of all d-stereoisomers in water were compared to experimental values with a good agreement.The relative binding free energies (ΔΔGbind) were calculated and, where available, compared to experimental values, approximated from Km values.The results suggest that a similar approach could be applied to study promiscuity of other enzymes.

View Article: PubMed Central - PubMed

Affiliation: Food Biotechnology Laboratory, Department of Food Science and Technology, University of Natural Resources and Life Sciences (BOKU), Vienna, Austria.

ABSTRACT
The flavoenzyme pyranose dehydrogenase (PDH) from the litter decomposing fungus Agaricus meleagris oxidizes many different carbohydrates occurring during lignin degradation. This promiscuous substrate specificity makes PDH a promising catalyst for bioelectrochemical applications. A generalized approach to simulate all 32 possible aldohexopyranoses in the course of one or a few molecular dynamics (MD) simulations is reported. Free energy calculations according to the one-step perturbation (OSP) method revealed the solvation free energies (ΔGsolv) of all 32 aldohexopyranoses in water, which have not yet been reported in the literature. The free energy difference between β- and α-anomers (ΔGβ-α) of all d-stereoisomers in water were compared to experimental values with a good agreement. Moreover, the free-energy differences (ΔG) of the 32 stereoisomers bound to PDH in two different poses were calculated from MD simulations. The relative binding free energies (ΔΔGbind) were calculated and, where available, compared to experimental values, approximated from Km values. The agreement was very good for one of the poses, in which the sugars are positioned in the active site for oxidation at C1 or C2. Distance analysis between hydrogens of the monosaccharide and the reactive N5-atom of the flavin adenine dinucleotide (FAD) revealed that oxidation is possible at HC1 or HC2 for pose A, and at HC3 or HC4 for pose B. Experimentally detected oxidation products could be rationalized for the majority of monosaccharides by combining ΔΔGbind and a reweighted distance analysis. Furthermore, several oxidation products were predicted for sugars that have not yet been tested experimentally, directing further analyses. This study rationalizes the relationship between binding free energies and substrate promiscuity in PDH, providing novel insights for its applicability in bioelectrochemistry. The results suggest that a similar approach could be applied to study promiscuity of other enzymes.

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Distributions of the improper dihedral 5 for the three 100 ns MD simulations of system SUGwater with changes to the topology according to Fig. 2.Coloring scheme: SUGa (black), SUGab (red), and SUGabc (blue).
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pcbi-1003995-g003: Distributions of the improper dihedral 5 for the three 100 ns MD simulations of system SUGwater with changes to the topology according to Fig. 2.Coloring scheme: SUGa (black), SUGab (red), and SUGabc (blue).

Mentions: To find a suitable reference state, which is crucial for reliable free energy calculations according to the OSP method, MD simulations of system SUGwater with changes to the topology according to SUGa, SUGab, and SUGabc were conducted. As a typical example, Fig. 3 shows the distributions of the improper dihedral angle 5 (ID5) centered on atom C5 for the three 100 ns MD simulations. For (black), ID5 is not evenly distributed and samples mostly the region around +30 degrees. (red) and (blue) both show more equal distributions, indicating that both stereo-configurations are equally sampled. To use a topology with minimal changes with respect to the real compounds, was selected as the most suitable reference state in water. Similarly, was used for the 1 µs SD simulation in vacuo (). In contrast, SUGa, SUGab, and SUGabc (Fig. 2) were all selected for simulations and analysis of system PDH-SUG, in order to sample as many stereoisomers as possible. Consequently, 12×50 ns MD simulations of system PDH-SUG were conducted: three different SUG topologies, two different SUG binding poses (pose A and pose B) and two independent simulations for each (md1 and md2). MD simulations of system PDH-SUG will be referred to as e.g. (pose A).


Pyranose dehydrogenase ligand promiscuity: a generalized approach to simulate monosaccharide solvation, binding, and product formation.

Graf MM, Zhixiong L, Bren U, Haltrich D, van Gunsteren WF, Oostenbrink C - PLoS Comput. Biol. (2014)

Distributions of the improper dihedral 5 for the three 100 ns MD simulations of system SUGwater with changes to the topology according to Fig. 2.Coloring scheme: SUGa (black), SUGab (red), and SUGabc (blue).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263366&req=5

pcbi-1003995-g003: Distributions of the improper dihedral 5 for the three 100 ns MD simulations of system SUGwater with changes to the topology according to Fig. 2.Coloring scheme: SUGa (black), SUGab (red), and SUGabc (blue).
Mentions: To find a suitable reference state, which is crucial for reliable free energy calculations according to the OSP method, MD simulations of system SUGwater with changes to the topology according to SUGa, SUGab, and SUGabc were conducted. As a typical example, Fig. 3 shows the distributions of the improper dihedral angle 5 (ID5) centered on atom C5 for the three 100 ns MD simulations. For (black), ID5 is not evenly distributed and samples mostly the region around +30 degrees. (red) and (blue) both show more equal distributions, indicating that both stereo-configurations are equally sampled. To use a topology with minimal changes with respect to the real compounds, was selected as the most suitable reference state in water. Similarly, was used for the 1 µs SD simulation in vacuo (). In contrast, SUGa, SUGab, and SUGabc (Fig. 2) were all selected for simulations and analysis of system PDH-SUG, in order to sample as many stereoisomers as possible. Consequently, 12×50 ns MD simulations of system PDH-SUG were conducted: three different SUG topologies, two different SUG binding poses (pose A and pose B) and two independent simulations for each (md1 and md2). MD simulations of system PDH-SUG will be referred to as e.g. (pose A).

Bottom Line: The free energy difference between β- and α-anomers (ΔGβ-α) of all d-stereoisomers in water were compared to experimental values with a good agreement.The relative binding free energies (ΔΔGbind) were calculated and, where available, compared to experimental values, approximated from Km values.The results suggest that a similar approach could be applied to study promiscuity of other enzymes.

View Article: PubMed Central - PubMed

Affiliation: Food Biotechnology Laboratory, Department of Food Science and Technology, University of Natural Resources and Life Sciences (BOKU), Vienna, Austria.

ABSTRACT
The flavoenzyme pyranose dehydrogenase (PDH) from the litter decomposing fungus Agaricus meleagris oxidizes many different carbohydrates occurring during lignin degradation. This promiscuous substrate specificity makes PDH a promising catalyst for bioelectrochemical applications. A generalized approach to simulate all 32 possible aldohexopyranoses in the course of one or a few molecular dynamics (MD) simulations is reported. Free energy calculations according to the one-step perturbation (OSP) method revealed the solvation free energies (ΔGsolv) of all 32 aldohexopyranoses in water, which have not yet been reported in the literature. The free energy difference between β- and α-anomers (ΔGβ-α) of all d-stereoisomers in water were compared to experimental values with a good agreement. Moreover, the free-energy differences (ΔG) of the 32 stereoisomers bound to PDH in two different poses were calculated from MD simulations. The relative binding free energies (ΔΔGbind) were calculated and, where available, compared to experimental values, approximated from Km values. The agreement was very good for one of the poses, in which the sugars are positioned in the active site for oxidation at C1 or C2. Distance analysis between hydrogens of the monosaccharide and the reactive N5-atom of the flavin adenine dinucleotide (FAD) revealed that oxidation is possible at HC1 or HC2 for pose A, and at HC3 or HC4 for pose B. Experimentally detected oxidation products could be rationalized for the majority of monosaccharides by combining ΔΔGbind and a reweighted distance analysis. Furthermore, several oxidation products were predicted for sugars that have not yet been tested experimentally, directing further analyses. This study rationalizes the relationship between binding free energies and substrate promiscuity in PDH, providing novel insights for its applicability in bioelectrochemistry. The results suggest that a similar approach could be applied to study promiscuity of other enzymes.

Show MeSH