The catalytic cycle of cytochrome P450: a fascinating choreography

Shaik, Sason; Dubey, Kshatresh Dutta

doi:10.1016/j.trechm.2021.09.004

Cited by 36 publications

(30 citation statements)

References 75 publications

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“…The catalytic cycle of P450 involves one-electron uptake, O 2 binding, a second electron uptake, and a proton uptake (left-side of Scheme a) to generate compound 0 (Cpd 0). ,− Since Cpd 0 is characterized in both the WT P450cam and the T252A mutant of P450cam, this species has usually been invoked as the second oxidant which performs substrate oxidation in P450 when the primary oxidant Cpd I could not be formed. − However, extensive computational and experimental evidence show that Cpd 0 is in fact not reactive in substrate oxidation. ,, …”

Section: Fenton-type H2o2 Activation Generates Cpd I In P450smentioning

confidence: 99%

How Do Metalloproteins Tame the Fenton Reaction and Utilize •OH Radicals in Constructive Manners?

et al. 2022

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Metrics & More Article Recommendations CONSPECTUS: This Account describes the manner whereby nature controls the Fenton-type reaction of O−O homolysis of hydrogen peroxide and harnesses it to carry out various useful oxidative transformations in metalloenzymes. H 2 O 2 acts as the cosubstrate for the heme-dependent peroxidases, P450BM3, P450 SPα , P450 BSβ , and the P450 decarboxylase OleT, as well as the nonheme enzymes HppE and the copper-dependent lytic polysaccharide monooxygenases (LPMOs). Whereas heme peroxidases use the Poulos-Kraut heterolytic mechanism for H 2 O 2 activation, some heme enzymes prefer the alternative Fentontype mechanism, which produces •OH radical intermediates. The fate of the •OH radical is controlled by the protein environment, using tight H-bonding networks around H 2 O 2 . The so-generated •OH radical is constrained by the surrounding H-bonding interactions, the orientation of which is targeted to perform Habstraction from the Fe(III)−OH group and thereby leading to the formation of the active species, called Compound I (Cpd I), Por +• Fe(IV)�O, which performs oxidation of the substrate. Alternatively, for the nonheme HppE enzyme, the O−O homolysis catalyzed by the resting state Fe(II) generates an Fe(III)−OH species that effectively constrains the •OH radical species by a tight H-bonding network. The so-formed H-bonded •OH radical acts directly as the oxidant, since it is oriented to perform H-abstraction from the C−H bond of the substrate (S)-2-HPP. The Fentontype H 2 O 2 activation is strongly suggested by computations to occur also in copper-dependent LPMOs and pMMO. In LPMOs, the Cu(I)-catalyzed O−O homolysis of the H 2 O 2 cosubstrate generates an •OH radical that abstracts a hydrogen atom from Cu(II)− OH and forms thereby the active species of the enzyme, Cu(II)-O•. Such Fenton-type O−O activation can be shared by both the O 2 -dependent activations of LPMOs and pMMOs, in which the O 2 cosubstrate may be reduced to H 2 O 2 by external reductants. Our studies show that, generally, the H 2 O 2 activation is highly dependent on the protein environment, as well as on the presence/absence of substrates. Since H 2 O 2 is a highly flexible and hydrophilic molecule, the absence of suitable substrates may lead to unproductive binding or even to the release of H 2 O 2 from the active site, as has been suggested in P450cam and LPMOs, whereas the presence of the substrate seems to play a role in steering a Fenton-type H 2 O 2 activation. In the absence of a substrate, the hydrophilic active site of P450BM3 disfavors the binding and activation of H 2 O 2 and protects thereby the enzyme from the damage by the Fenton reaction. Due to the distinct coordination and reaction environment, the Fenton-type H 2 O 2 activation mechanism by enzymes differs from the reaction in synthetic systems. In nonenzymatic reactions, the H-bonding networks are quite dynamic and flexible and the reactivity of H 2 O 2 is not strategically constrained as in the enzymatic environment. As such, our Account describes...

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Section: Fenton-type H2o2 Activation Generates Cpd I In P450smentioning

confidence: 99%

How Do Metalloproteins Tame the Fenton Reaction and Utilize •OH Radicals in Constructive Manners?

et al. 2022

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“…Cytochromes P450 are nature’s efficient nanomachines that catalyze a plethora of reactions in all organisms. − These enzymes are phenomenally versatile; they are responsible for detoxification of foreign compounds (so-called xenobiotics), and among other processes, they can perform, for example, sulfoxidation, C–H/CC activations, carbene- and nitrile-insertion, as well as C–C bond making and cleavage. − Such a synthetic versatility makes P450 a favorite model for bio-engineering and evolutionary studies. − The paradigmatic catalytic cycle of P450 starts by the entry of a substrate, which triggers concomitant delivery of two electrons (from a reducing partner) and two protons, finally leading to the formation of the porphyrin-radical-cation oxoiron species, the so-called compound I (Cpd I), which in turn oxidizes the substrate. − This mechanistic paradigm has prevailed until the discovery of P450 peroxygenases which utilize hydrogen peroxide to shunt the catalytic cycle and thereby bypasses the necessity of a reducing partner …”

Section: Introductionmentioning

confidence: 99%

Local Electric Fields Dictate Function: The Different Product Selectivities Observed for Fatty Acid Oxidation by Two Deceptively Very Similar P450-Peroxygenases OleT and BSβ

Yadav

Shaik

Siddiqui

et al. 2022

J. Chem. Inf. Model.

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Cytochrome P450 peroxygenases use hydrogen peroxide to hydroxylate long-chain fatty acids by bypassing the use of O 2 and a redox partner. Among the peroxygenases, P450 OleT uniquely performs decarboxylation of fatty acids and production of terminal olefins. This route taken by P450 OleT is intriguing, and its importance is augmented by the practical importance of olefin production. As such, this mechanistic choice merits elucidation. To address this puzzle, we use hybrid QM/ MM calculations and MD simulations for the OleT enzyme as well as for the structurally analogous enzyme, P450 BSβ . The study of P450 OleT reveals that the protonated His85 in the wild-type P450 OleT plays a crucial role in steering decarboxylation activity by stabilizing the corresponding hydroxoiron(IV) intermediate (Cpd II). In contrast, for P450 BSβ in which Q85 replaces H85, the respective Cpd II species is unstable and it reacts readily with the substrate radical by rebound, producing hydroxylation products. As shown, this single-site difference creates in P450 OleT a local electric field (LEF), which is significantly higher than that in P450 BSβ . In turn, these LEF differences are responsible for the different stabilities of the respective Cpd II/radical intermediates and hence for different functions of the two enzymes. P450 BSβ uses the common rebound mechanism and leads to hydroxylation, whereas P450 OleT proceeds via decarboxylation and generates terminal olefins. Olefin production projects the power of a single residue to alter the LEF and the enzyme's function.

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“…Cytochrome P450 is nature’s ultimate enzyme that can catalyze a plethora of chemical reactions. − It is biological machinery that works in a well-organized catalytic cycle using molecular oxygen, water molecules, and electrons as fuel for the efficient oxidation of several compounds of commercial and medicinal importance. − Due to the catalytic versatility of P450 enzymes, it is no surprise that cytochrome P450 is the most widely used scaffold for bioengineering of new catalytic functions. − Recently, three members of the CYP450 superfamily, P450 OleT from the peroxygenase family, , P450 GcoA from CYP255A2, − and AgcA from the CYP255A1 family, were found to catalyze reactions of potential biofuel importance . P450 GcoA , in particular, gained special attention as it catalyzes the downstream degradation of lignin, which can potentially be used to produce a drop-in biofuel …”

Section: Introductionmentioning

confidence: 99%

Single-Site Mutation Induces Water-Mediated Promiscuity in Lignin Breaking Cytochrome P450_GcoA

et al. 2022

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Cytochrome P450 GcoA is an enzyme that catalyzes the guaiacol unit of lignin during the lignin breakdown via an aryl- O -demethylation reaction. This reaction is intriguing and is of commercial importance for its potential applications in the production of biofuel and plastic from biomass feedstock. Recently, the F169A mutation in P450 GcoA elicits a promiscuous activity for syringol while maintaining the native activity for guaiacol. Using comprehensive MD simulations and hybrid QM/MM calculations, we address, herein, the origin of promiscuity in P450 GcoA and its relevance to the specific activity toward lignin-derived substrates. Our study shows a crucial role of an aromatic dyad of F169 and F395 by regulating the water access to the catalytic center. The F169A mutation opens a water aqueduct and hence increases the native activity for G-lignin. We show that syringol binds very tightly to the WT enzyme, which blocks the conformational rearrangement needed for the second step of O-demethylation. The F169A creates an extra room favoring the conformational rearrangement in the 3-methoxycatechol (3MC) and second dose of the dioxygen insertion. Therefore, using MD simulations and complemented by thorough QM/MM calculations, our study shows how a single-site mutation rearchitects active site engineering for promiscuous syringol activity.

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The catalytic cycle of cytochrome P450: a fascinating choreography

Cited by 36 publications

References 75 publications

How Do Metalloproteins Tame the Fenton Reaction and Utilize •OH Radicals in Constructive Manners?

How Do Metalloproteins Tame the Fenton Reaction and Utilize •OH Radicals in Constructive Manners?

Local Electric Fields Dictate Function: The Different Product Selectivities Observed for Fatty Acid Oxidation by Two Deceptively Very Similar P450-Peroxygenases OleT and BSβ

Single-Site Mutation Induces Water-Mediated Promiscuity in Lignin Breaking Cytochrome P450_GcoA

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The catalytic cycle of cytochrome P450: a fascinating choreography

Cited by 36 publications

References 75 publications

How Do Metalloproteins Tame the Fenton Reaction and Utilize •OH Radicals in Constructive Manners?

How Do Metalloproteins Tame the Fenton Reaction and Utilize •OH Radicals in Constructive Manners?

Local Electric Fields Dictate Function: The Different Product Selectivities Observed for Fatty Acid Oxidation by Two Deceptively Very Similar P450-Peroxygenases OleT and BSβ

Single-Site Mutation Induces Water-Mediated Promiscuity in Lignin Breaking Cytochrome P450GcoA

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Single-Site Mutation Induces Water-Mediated Promiscuity in Lignin Breaking Cytochrome P450_GcoA