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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.


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(a) Chemical structuresof 1-meA and 1-deazameA. (b, c, d) 1-DeazameA-relatedintermediates. Carbon atoms are colored in gray, hydrogen in white,nitrogen in blue, oxygen in red, iron in purple, and boundary carbonatoms for pseudo-bond in cyan.
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fig7: (a) Chemical structuresof 1-meA and 1-deazameA. (b, c, d) 1-DeazameA-relatedintermediates. Carbon atoms are colored in gray, hydrogen in white,nitrogen in blue, oxygen in red, iron in purple, and boundary carbonatoms for pseudo-bond in cyan.

Mentions: In order to investigate the effect from thepositive charge onthe nitrogen (N1) of 1-meA, we changed 1-meA to 1-deazameA by replacingN1 with C (see Figure 7a) in the optimizedzwitterion I2OH (bound to the iron) and I5OH (unbound from the iron) structures and carried out geometry optimizations.A stable structure for the I2OH analog (Figure 7b) for 1-deazameA was obtained on the PES. Conversely,in the case of the I5OH analog for 1-deazameA, the −CH2O– group abstracts a proton spontaneouslyfrom the neighboring water that is coordinated to the iron (Figure 7c). This finding is consistent with previous resultsfor 3-deazameC.3 Additionally, differentfrom 1-meA, we are able to obtain the I3OH analog (Figure 7d) for 1-deazmaeA, although its energy is 8.0 kcal/molhigher than for the I2OH analog. These optimized structuresindicate that the proton accepting ability of iron-bound −CH2O– group in 1-deazameA is weaker than whenthis group is not bound to the iron because of the stabilization effectsfrom the metal. The pKa of iron-unbound−CH2OH in 1-deazameA is larger than the value ofH2O coordinated to the iron, and hence larger than 1meA.Since the proton transfer is a necessary step for the repair processand might become the rate-limiting step as discussed in the next section,the pKa difference of −CH2OH may partly explain why 1-meA and 3-meC, which bear a positivecharge, are preferred by AlkB over 3-meT and 1-meG, which are chargeneutral alkylated DNA bases.3,8


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)

(a) Chemical structuresof 1-meA and 1-deazameA. (b, c, d) 1-DeazameA-relatedintermediates. Carbon atoms are colored in gray, hydrogen in white,nitrogen in blue, oxygen in red, iron in purple, and boundary carbonatoms for pseudo-bond in cyan.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4230374&req=5

fig7: (a) Chemical structuresof 1-meA and 1-deazameA. (b, c, d) 1-DeazameA-relatedintermediates. Carbon atoms are colored in gray, hydrogen in white,nitrogen in blue, oxygen in red, iron in purple, and boundary carbonatoms for pseudo-bond in cyan.
Mentions: In order to investigate the effect from thepositive charge onthe nitrogen (N1) of 1-meA, we changed 1-meA to 1-deazameA by replacingN1 with C (see Figure 7a) in the optimizedzwitterion I2OH (bound to the iron) and I5OH (unbound from the iron) structures and carried out geometry optimizations.A stable structure for the I2OH analog (Figure 7b) for 1-deazameA was obtained on the PES. Conversely,in the case of the I5OH analog for 1-deazameA, the −CH2O– group abstracts a proton spontaneouslyfrom the neighboring water that is coordinated to the iron (Figure 7c). This finding is consistent with previous resultsfor 3-deazameC.3 Additionally, differentfrom 1-meA, we are able to obtain the I3OH analog (Figure 7d) for 1-deazmaeA, although its energy is 8.0 kcal/molhigher than for the I2OH analog. These optimized structuresindicate that the proton accepting ability of iron-bound −CH2O– group in 1-deazameA is weaker than whenthis group is not bound to the iron because of the stabilization effectsfrom the metal. The pKa of iron-unbound−CH2OH in 1-deazameA is larger than the value ofH2O coordinated to the iron, and hence larger than 1meA.Since the proton transfer is a necessary step for the repair processand might become the rate-limiting step as discussed in the next section,the pKa difference of −CH2OH may partly explain why 1-meA and 3-meC, which bear a positivecharge, are preferred by AlkB over 3-meT and 1-meG, which are chargeneutral alkylated DNA bases.3,8

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