Limits...
Carbinolamine formation and dehydration in a DNA repair enzyme active site.

Dodson ML, Walker RC, Lloyd RS - PLoS ONE (2012)

Bottom Line: We demonstrated feasible pathways involving water, as well as those independent of water participation.Water-independent unforced proton transfer from the protonated active site glutamate carboxyl to the unprotonated N-terminal amine was also observed.Imine carbinolamine formation was characterized using steered QM/MM molecular dynamics.

View Article: PubMed Central - PubMed

Affiliation: Active Site Dynamics LLC, Houston, Texas, United States of America. mldodson@comcast.net

ABSTRACT
In order to suggest detailed mechanistic hypotheses for the formation and dehydration of a key carbinolamine intermediate in the T4 pyrimidine dimer glycosylase (T4PDG) reaction, we have investigated these reactions using steered molecular dynamics with a coupled quantum mechanics-molecular mechanics potential (QM/MM). We carried out simulations of DNA abasic site carbinolamine formation with and without a water molecule restrained to remain within the active site quantum region. We recovered potentials of mean force (PMF) from thirty replicate reaction trajectories using Jarzynski averaging. We demonstrated feasible pathways involving water, as well as those independent of water participation. The water-independent enzyme-catalyzed reaction had a bias-corrected Jarzynski-average barrier height of approximately (6.5 kcal mol(-1) (27.2 kJ mol(-1)) for the carbinolamine formation reaction and 44.5 kcal mol(-1) (186 kJ mol(-1)) for the reverse reaction at this level of representation. When the proton transfer was facilitated with an intrinsic quantum water, the barrier height was approximately 15 kcal mol(-1) (62.8 kJ mol(-1)) in the forward (formation) reaction and 19 kcal mol(-1) (79.5 kJ mol(-1)) for the reverse. In addition, two modes of unsteered (free dynamics) carbinolamine dehydration were observed: in one, the quantum water participated as an intermediate proton transfer species, and in the other, the active site protonated glutamate hydrogen was directly transferred to the carbinolamine oxygen. Water-independent unforced proton transfer from the protonated active site glutamate carboxyl to the unprotonated N-terminal amine was also observed. In summary, complex proton transfer events, some involving water intermediates, were studied in QM/MM simulations of T4PDG bound to a DNA abasic site. Imine carbinolamine formation was characterized using steered QM/MM molecular dynamics. Dehydration of the carbinolamine intermediate to form the final imine product was observed in free, unsteered, QM/MM dynamics simulations, as was unforced acid-base transfer between the active site carboxylate and the N-terminal amine.

Show MeSH

Related in: MedlinePlus

Stereogram of a typical initial quantum region for carbinolamine formation mediated by an active site water.The large purple “atoms” are the link hydrogens, proxies for the classical mechanics sites covalently bonded to the quantum region. The atomic coordinates for this figure were taken directly from one of the Protein Data Bank (PDB) files for the quantum atoms written at the beginning of each QM/MM run. Molecular graphics in this and other similar figures were prepared with MOLSCRIPT [37] and Raster3D [38].
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3285167&req=5

pone-0031377-g002: Stereogram of a typical initial quantum region for carbinolamine formation mediated by an active site water.The large purple “atoms” are the link hydrogens, proxies for the classical mechanics sites covalently bonded to the quantum region. The atomic coordinates for this figure were taken directly from one of the Protein Data Bank (PDB) files for the quantum atoms written at the beginning of each QM/MM run. Molecular graphics in this and other similar figures were prepared with MOLSCRIPT [37] and Raster3D [38].

Mentions: Fig. 2 illustrates the initial active site quantum region structure for the SMD simulation analyzed in Fig. 3. The atom names are those referred to in the collective variable definition in Fig. 3, Panel B. Several distances are also defined: , , and , where is the distance between atoms and . The starting value of the SMD CV was its value at the end of the 1 ns QM/MM relaxation run (8.92 Å). The ending value (3.52 Å) was taken to be the sum of the bond lengths: . In this and similar simulation setups, the quantum water molecule(s) were restrained to be within a flat well potential spherical volume around with weak harmonic restraint walls. Further details are given in the file S3.


Carbinolamine formation and dehydration in a DNA repair enzyme active site.

Dodson ML, Walker RC, Lloyd RS - PLoS ONE (2012)

Stereogram of a typical initial quantum region for carbinolamine formation mediated by an active site water.The large purple “atoms” are the link hydrogens, proxies for the classical mechanics sites covalently bonded to the quantum region. The atomic coordinates for this figure were taken directly from one of the Protein Data Bank (PDB) files for the quantum atoms written at the beginning of each QM/MM run. Molecular graphics in this and other similar figures were prepared with MOLSCRIPT [37] and Raster3D [38].
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3285167&req=5

pone-0031377-g002: Stereogram of a typical initial quantum region for carbinolamine formation mediated by an active site water.The large purple “atoms” are the link hydrogens, proxies for the classical mechanics sites covalently bonded to the quantum region. The atomic coordinates for this figure were taken directly from one of the Protein Data Bank (PDB) files for the quantum atoms written at the beginning of each QM/MM run. Molecular graphics in this and other similar figures were prepared with MOLSCRIPT [37] and Raster3D [38].
Mentions: Fig. 2 illustrates the initial active site quantum region structure for the SMD simulation analyzed in Fig. 3. The atom names are those referred to in the collective variable definition in Fig. 3, Panel B. Several distances are also defined: , , and , where is the distance between atoms and . The starting value of the SMD CV was its value at the end of the 1 ns QM/MM relaxation run (8.92 Å). The ending value (3.52 Å) was taken to be the sum of the bond lengths: . In this and similar simulation setups, the quantum water molecule(s) were restrained to be within a flat well potential spherical volume around with weak harmonic restraint walls. Further details are given in the file S3.

Bottom Line: We demonstrated feasible pathways involving water, as well as those independent of water participation.Water-independent unforced proton transfer from the protonated active site glutamate carboxyl to the unprotonated N-terminal amine was also observed.Imine carbinolamine formation was characterized using steered QM/MM molecular dynamics.

View Article: PubMed Central - PubMed

Affiliation: Active Site Dynamics LLC, Houston, Texas, United States of America. mldodson@comcast.net

ABSTRACT
In order to suggest detailed mechanistic hypotheses for the formation and dehydration of a key carbinolamine intermediate in the T4 pyrimidine dimer glycosylase (T4PDG) reaction, we have investigated these reactions using steered molecular dynamics with a coupled quantum mechanics-molecular mechanics potential (QM/MM). We carried out simulations of DNA abasic site carbinolamine formation with and without a water molecule restrained to remain within the active site quantum region. We recovered potentials of mean force (PMF) from thirty replicate reaction trajectories using Jarzynski averaging. We demonstrated feasible pathways involving water, as well as those independent of water participation. The water-independent enzyme-catalyzed reaction had a bias-corrected Jarzynski-average barrier height of approximately (6.5 kcal mol(-1) (27.2 kJ mol(-1)) for the carbinolamine formation reaction and 44.5 kcal mol(-1) (186 kJ mol(-1)) for the reverse reaction at this level of representation. When the proton transfer was facilitated with an intrinsic quantum water, the barrier height was approximately 15 kcal mol(-1) (62.8 kJ mol(-1)) in the forward (formation) reaction and 19 kcal mol(-1) (79.5 kJ mol(-1)) for the reverse. In addition, two modes of unsteered (free dynamics) carbinolamine dehydration were observed: in one, the quantum water participated as an intermediate proton transfer species, and in the other, the active site protonated glutamate hydrogen was directly transferred to the carbinolamine oxygen. Water-independent unforced proton transfer from the protonated active site glutamate carboxyl to the unprotonated N-terminal amine was also observed. In summary, complex proton transfer events, some involving water intermediates, were studied in QM/MM simulations of T4PDG bound to a DNA abasic site. Imine carbinolamine formation was characterized using steered QM/MM molecular dynamics. Dehydration of the carbinolamine intermediate to form the final imine product was observed in free, unsteered, QM/MM dynamics simulations, as was unforced acid-base transfer between the active site carboxylate and the N-terminal amine.

Show MeSH
Related in: MedlinePlus