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Alternative Pathway for the Reaction Catalyzed by DNA Dealkylase AlkB from Ab Initio QM/MM Calculations.

Fang D, Cisneros GA - J Chem Theory Comput (2014)

Bottom Line: The new OH rebound step is coupled with a proton transfer to the OH(-) ligand and results in a novel zwitterion intermediate.The consistency between our theoretical results and experimental findings is discussed.This study provides new insights into the oxidative repair mechanism of DNA repair by nonheme Fe(II) and α-ketoglutarate (α-KG) dependent dioxygenases and a possible explanation for the substrate preference of AlkB.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Wayne State University , Detroit, Michigan 48202, United States.

ABSTRACT
AlkB is the title enzyme of a family of DNA dealkylases that catalyze the direct oxidative dealkylation of nucleobases. The conventional mechanism for the dealkylation of N(1)-methyl adenine (1-meA) catalyzed by AlkB after the formation of Fe(IV)-oxo is comprised by a reorientation of the oxo moiety, hydrogen abstraction, OH rebound from the Fe atom to the methyl adduct, and the dissociation of the resulting methoxide to obtain the repaired adenine base and formaldehyde. An alternative pathway with hydroxide as a ligand bound to the iron atom is proposed and investigated by QM/MM simulations. The results show OH(-) has a small impact on the barriers for the hydrogen abstraction and OH rebound steps. The effects of the enzyme and the OH(-) ligand on the hydrogen abstraction by the Fe(IV)-oxo moiety are discussed in detail. The new OH rebound step is coupled with a proton transfer to the OH(-) ligand and results in a novel zwitterion intermediate. This zwitterion structure can also be characterized as Fe-O-C complex and facilitates the formation of formaldehyde. In contrast, for the pathway with H2O bound to iron, the hydroxyl product of the OH rebound step first needs to unbind from the metal center before transferring a proton to Glu136 or other residue/substrate. The consistency between our theoretical results and experimental findings is discussed. This study provides new insights into the oxidative repair mechanism of DNA repair by nonheme Fe(II) and α-ketoglutarate (α-KG) dependent dioxygenases and a possible explanation for the substrate preference of AlkB.

No MeSH data available.


Related in: MedlinePlus

Relative energies (kcal/mol) for the structures alongthe minimumenergy path (MEP) for the proton transferred to Glu136 in the quintetstate for the H2O pathway. (Arg210, Glu136, and a bridgingwater were added to the QM subsystem for these structures). Carbonatoms are colored in gray, hydrogen in white, nitrogen in blue, oxygenin red, iron in purple, and boundary carbon atoms for pseudo-bondin cyan.
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fig9: Relative energies (kcal/mol) for the structures alongthe minimumenergy path (MEP) for the proton transferred to Glu136 in the quintetstate for the H2O pathway. (Arg210, Glu136, and a bridgingwater were added to the QM subsystem for these structures). Carbonatoms are colored in gray, hydrogen in white, nitrogen in blue, oxygenin red, iron in purple, and boundary carbon atoms for pseudo-bondin cyan.

Mentions: It is also possiblethat the proton is transferred to a neighboringresidue instead of being transferred to Asp133. As shown in Figure 1, the nearest residue to the active site is Glu136,but its distance from the 1-meA suggests the proton transfer wouldlikely have to occur via a water bridge. Before the proton transfer,the structure rearranges from I3Glu136 to I3′Glu136. (Figure 9). In I3Glu136, the −CH3OH moiety forms a hydrogen bond withAsp133 while −CH3OH forms a hydrogen bond with thebridging water in I3′Glu136. To check the existenceof a zwitterion structure, we carried out the optimization of I4Glu136 assuming the proton transfer from −CH3OH to Glu136 with a water molecule as the bridge. However, duringthe optimization, the structure changes back to I3′Glu136 with the proton being spontaneously transferred back. This indicatesthat the pKa of −CH3OH is also larger than Glu136H. In other words, if the proton istransferred to Glu136, it has to be coupled with the C–N bondbreaking. The calculated barrier for TSI3′Pun-Glu136 for the concerted pathway from I3′Glu136 to PGlu136 is 7.4 kcal/mol, which is much lower than the barrierfor the proton being transferred to Asp133. This suggests that Glu136may be the final proton acceptor when no better acceptor available.The proton acceptor role of Glu136 may partly account for the decreasedactivity of AlkB in repairing 1-meA when Glu136 is mutated to a leucine.39


Alternative Pathway for the Reaction Catalyzed by DNA Dealkylase AlkB from Ab Initio QM/MM Calculations.

Fang D, Cisneros GA - J Chem Theory Comput (2014)

Relative energies (kcal/mol) for the structures alongthe minimumenergy path (MEP) for the proton transferred to Glu136 in the quintetstate for the H2O pathway. (Arg210, Glu136, and a bridgingwater were added to the QM subsystem for these structures). Carbonatoms are colored in gray, hydrogen in white, nitrogen in blue, oxygenin red, iron in purple, and boundary carbon atoms for pseudo-bondin cyan.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4230374&req=5

fig9: Relative energies (kcal/mol) for the structures alongthe minimumenergy path (MEP) for the proton transferred to Glu136 in the quintetstate for the H2O pathway. (Arg210, Glu136, and a bridgingwater were added to the QM subsystem for these structures). Carbonatoms are colored in gray, hydrogen in white, nitrogen in blue, oxygenin red, iron in purple, and boundary carbon atoms for pseudo-bondin cyan.
Mentions: It is also possiblethat the proton is transferred to a neighboringresidue instead of being transferred to Asp133. As shown in Figure 1, the nearest residue to the active site is Glu136,but its distance from the 1-meA suggests the proton transfer wouldlikely have to occur via a water bridge. Before the proton transfer,the structure rearranges from I3Glu136 to I3′Glu136. (Figure 9). In I3Glu136, the −CH3OH moiety forms a hydrogen bond withAsp133 while −CH3OH forms a hydrogen bond with thebridging water in I3′Glu136. To check the existenceof a zwitterion structure, we carried out the optimization of I4Glu136 assuming the proton transfer from −CH3OH to Glu136 with a water molecule as the bridge. However, duringthe optimization, the structure changes back to I3′Glu136 with the proton being spontaneously transferred back. This indicatesthat the pKa of −CH3OH is also larger than Glu136H. In other words, if the proton istransferred to Glu136, it has to be coupled with the C–N bondbreaking. The calculated barrier for TSI3′Pun-Glu136 for the concerted pathway from I3′Glu136 to PGlu136 is 7.4 kcal/mol, which is much lower than the barrierfor the proton being transferred to Asp133. This suggests that Glu136may be the final proton acceptor when no better acceptor available.The proton acceptor role of Glu136 may partly account for the decreasedactivity of AlkB in repairing 1-meA when Glu136 is mutated to a leucine.39

Bottom Line: The new OH rebound step is coupled with a proton transfer to the OH(-) ligand and results in a novel zwitterion intermediate.The consistency between our theoretical results and experimental findings is discussed.This study provides new insights into the oxidative repair mechanism of DNA repair by nonheme Fe(II) and α-ketoglutarate (α-KG) dependent dioxygenases and a possible explanation for the substrate preference of AlkB.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Wayne State University , Detroit, Michigan 48202, United States.

ABSTRACT
AlkB is the title enzyme of a family of DNA dealkylases that catalyze the direct oxidative dealkylation of nucleobases. The conventional mechanism for the dealkylation of N(1)-methyl adenine (1-meA) catalyzed by AlkB after the formation of Fe(IV)-oxo is comprised by a reorientation of the oxo moiety, hydrogen abstraction, OH rebound from the Fe atom to the methyl adduct, and the dissociation of the resulting methoxide to obtain the repaired adenine base and formaldehyde. An alternative pathway with hydroxide as a ligand bound to the iron atom is proposed and investigated by QM/MM simulations. The results show OH(-) has a small impact on the barriers for the hydrogen abstraction and OH rebound steps. The effects of the enzyme and the OH(-) ligand on the hydrogen abstraction by the Fe(IV)-oxo moiety are discussed in detail. The new OH rebound step is coupled with a proton transfer to the OH(-) ligand and results in a novel zwitterion intermediate. This zwitterion structure can also be characterized as Fe-O-C complex and facilitates the formation of formaldehyde. In contrast, for the pathway with H2O bound to iron, the hydroxyl product of the OH rebound step first needs to unbind from the metal center before transferring a proton to Glu136 or other residue/substrate. The consistency between our theoretical results and experimental findings is discussed. This study provides new insights into the oxidative repair mechanism of DNA repair by nonheme Fe(II) and α-ketoglutarate (α-KG) dependent dioxygenases and a possible explanation for the substrate preference of AlkB.

No MeSH data available.


Related in: MedlinePlus