QM/MM Study of the Reaction Mechanism of the Dehydratase Domain from Mammalian Fatty Acid Synthase

Medina, Fabiola E.; Neves, Rui P. P.; Ramos, Maria João; Fernandes, Pedro A.

doi:10.1021/acscatal.8b02616

Cited by 39 publications

(61 citation statements)

References 75 publications

(139 reference statements)

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“…The oxyanion hole stabilization has also been quantified in other studies that focused on enzymes such as subtilisin, thiolase, ketosteroid isomerase, dehydratase, trypsin and transferase . These studies reported an oxyanion hole‐mediated stabilization of transition‐states of 4–6 kcal ⋅ mol −1 , which is lower than the 7–11 kcal ⋅ mol −1 presented in the current work.…”

Section: Resultsmentioning

confidence: 43%

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Human Fatty Acid Synthase: A Computational Study of the Transfer of the Acyl Moieties from MAT to the ACP Domain

et al. 2019

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Human Fatty Acid Synthase (hFAS) is a multidomain enzyme responsible for the biosynthesis of saturated fatty acids, which are crucial for numerous biological processes. During the biosynthetic reaction, hFAS incorporates acetyl and malonyl moieties through the action of its malonyl‐acetyl transferase (MAT) domain. These precursor molecules are transferred from CoA to the acyl‐carrier protein (ACP) domain by a two‐stage ping‐pong catalytic mechanism. The first stage, during which the acyl moieties are relocated from CoA to the MAT domain, has been elucidated in a previous work of ours. In this study, we employ QM/MM calculations at the DLPNO‐CCSD(T)/CBS:AMBER level of theory to understand the catalytic machinery behind the transfer of the acyl moieties from MAT to the ACP domain. The results show that the acyl transfer to the ACP's phosphopantetheine (PNS) occurs in two sequential events: Step 3) asynchronous concerted reaction that combines the deprotonation of the PNS's thiol group and the nucleophilic attack on the acyl‐Ser581 ester carbon; Step 4) tetrahedral intermediate breakdown and regeneration of the catalytic Ser581/His683 dyad. The energy barriers of these two steps are inferior to those reported for the first catalytic stage, suggesting that the rate‐limiting step of the MAT mechanism lies on Stage 1, particularly on the concerted nucleophilic attack that starts the reaction. The oxyanion hole, formed by the backbone amines of Met499 and Leu582, was reported to transiently stabilize the tetrahedral intermediate and to play a favorable catalytic effect on the overall MAT reaction, particularly through the reduction of the energetic span. The knowledge gathered in this work complements previous experimental and theoretical studies and instigates further investigations that explore the therapeutic potential of hFAS.

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Section: Resultsmentioning

confidence: 43%

“…Indeed, this 7–11 kcal ⋅ mol −1 energy range lays close to the 11.1 kcal ⋅ mol −1 value reported by Medina et al . after performing a double replacement protocol …”

Section: Resultsmentioning

confidence: 99%

Human Fatty Acid Synthase: A Computational Study of the Transfer of the Acyl Moieties from MAT to the ACP Domain

et al. 2019

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“…In order to investigate the catalytic mechanism, hybrid QM/MM procedure was applied as previously reported in similar systems [ 10 , 11 , 12 , 13 , 14 , 15 , 16 ]. The QM region includes Cys18, His22 of the seven chains and Glu25 faced into the hydrophobic channel of the CC-Hept-Cys-His-Glu and three water molecules (w1, w2 and w3) engaged in a hydrogen bonds network (see Figure 4 ) both with each other and with the catalytic triad ( Figure S6 ).…”

Section: Resultsmentioning

confidence: 99%

“…In order to quantify entropic contributions, the Grimme’s procedure [ 50 ] was employed as reported in other works [ 14 , 15 , 16 ]. According to this method, only vibrational temperatures higher than 100 K are taken into account to calculate TΔS contribution using the harmonic oscillator/rigid rotor/particle in a box formalism.…”

Section: Methodsmentioning

confidence: 99%

On the Catalytic Activity of the Engineered Coiled-Coil Heptamer Mimicking the Hydrolase Enzymes: Insights from a Computational Study

Prejanò

Romeo

Russo

et al. 2020

IJMS

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Recently major advances were gained on the designed proteins aimed to generate biomolecular mimics of proteases. Although such enzyme-like catalysts must still suffer refinements for improving the catalytic activity, at the moment, they represent a good example of artificial enzymes to be tested in different fields. Herein, a de novo designed homo-heptameric peptide assembly (CC-Hept) where the esterase activity towards p-nitro-phenylacetate was obtained for introduction of the catalytic triad (Cys-His-Glu) into the hydrophobic matrix, is the object of the present combined molecular dynamics and quantum mechanics/molecular mechanics investigation. Constant pH Molecular Dynamics simulations on the apoform of CC-Hept suggested that the Cys residues are present in the protonated form. Molecular dynamics (MD) simulations of the enzyme–substrate complex evidenced the attitude of the enzyme-like system to retain water molecules, necessary in the hydrolytic reaction, in correspondence of the active site, represented by the Cys-His-Glu triad on each of the seven chains, without significant structural perturbations. A detailed reaction mechanism of esterase activity of CC-Hept-Cys-His-Glu was investigated on the basis of the quantum mechanics/molecular mechanics calculations employing a large quantum mechanical (QM) region of the active site. The proposed mechanism is consistent with available esterases kinetics and structural data. The roles of the active site residues were also evaluated. The deacylation phase emerged as the rate-determining step, in agreement with esterase activity of other natural proteases.

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“…Only vibrational temperatures larger than 100 K have been accounted for, during the calculation of the entropy contribution, according to a validated procedure, [49] already successfully used in other works. [50][51][52] The final energy profiles were obtained adding the Zero Point Energy, the empirical D3-dispersion correction [47] and entropy contributions, as reported in Table S1.…”

Section: Methodsmentioning

confidence: 99%

The Catalytic Mechanism of Human Transketolase

et al. 2019

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We have computationally determined the catalytic mechanism of human transketolase (hTK) using a cluster model approach and density functional theory calculations. We were able to determine all the relevant structures, bringing solid evidences to the proposed experimental mechanism, and to add important detail to the structure of the transition states and the energy profile associated with catalysis. Furthermore, we have established the existence of a crucial intermediate of the catalytic cycle, in agreement with experiments. The calculated data brought new insights to hTK's catalytic mechanism, providing free-energy values for the chemical reaction, as well as adding atomistic detail to the experimental mechanism.[a] Dr.

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QM/MM Study of the Reaction Mechanism of the Dehydratase Domain from Mammalian Fatty Acid Synthase

Cited by 39 publications

References 75 publications

Human Fatty Acid Synthase: A Computational Study of the Transfer of the Acyl Moieties from MAT to the ACP Domain

Human Fatty Acid Synthase: A Computational Study of the Transfer of the Acyl Moieties from MAT to the ACP Domain

On the Catalytic Activity of the Engineered Coiled-Coil Heptamer Mimicking the Hydrolase Enzymes: Insights from a Computational Study

The Catalytic Mechanism of Human Transketolase

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