New Features in the Catalytic Cycle of Cytochrome P450 during the Formation of Compound I from Compound 0

2005 ◽  
Vol 109 (42) ◽  
pp. 19946-19951 ◽  
Author(s):  
Devesh Kumar ◽  
Hajime Hirao ◽  
Sam P. de Visser ◽  
Jingjing Zheng ◽  
Dongqi Wang ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Catalytic Cycle ◽  
Compound I

Density functional theory (DFT) and combined quantum mechanical/molecular mechanics (QM/MM) studies on the oxygen activation step in nitric oxide synthase enzymes

2009 ◽  
Vol 37 (2) ◽  
pp. 373-377 ◽  
Author(s):  
Sam P. de Visser
Keyword(s):  
Nitric Oxide ◽  
Density Functional Theory ◽  
Cytochrome P450 ◽  
Density Functional ◽  
Catalytic Cycle ◽  
Theoretical Studies ◽  
Cytochrome P450 Enzymes ◽  
Compound I ◽  
Functional Theory ◽  
Oxide Synthase

In this review paper, we will give an overview of recent theoretical studies on the catalytic cycle(s) of NOS (nitric oxide synthase) enzymes and in particular on the later stages of these cycles where experimental work is difficult due to the short lifetime of intermediates. NOS enzymes are vital for human health and are involved in the biosynthesis of toxic nitric oxide. Despite many experimental efforts in the field, the catalytic cycle of this important enzyme is still surrounded by many unknowns and controversies. Our theoretical studies were focused on the grey zones of the catalytic cycle, where intermediates are short-lived and experimental detection is impossible. Thus combined QM/MM (quantum mechanics/molecular mechanics) as well as DFT (density functional theory) studies on NOS enzymes and active site models have established a novel mechanism of oxygen activation and the conversion of L-arginine into Nω-hydroxo-arginine. Although NOS enzymes show many structural similarities to cytochrome P450 enzymes, it has long been anticipated that therefore they should have a similar catalytic cycle where molecular oxygen binds to a haem centre and is converted into an Fe(IV)-oxo haem(+•) active species (Compound I). Compound I, however, is elusive in the cytochrome P450s as well as in NOS enzymes, but indirect experimental evidence on cytochrome P450 systems combined with theoretical modelling have shown it to be the oxidant responsible for hydroxylation reactions in cytochrome P450 enzymes. By contrast, in the first catalytic cycle of NOS it has been shown that Compound I is first reduced to Compound II before the hydroxylation of arginine. Furthermore, substrate arginine in NOS enzymes appears to have a dual function, namely first as a proton donor in the catalytic cycle to convert the ferric-superoxo into a ferric-hydroperoxo complex and secondly as the substrate that is hydroxylated in the process leading to Nω-hydroxo-arginine.

One small step for cytochrome P450 in its catalytic cycle, one giant leap for enzymology

2019 ◽  
Vol 23 (04n05) ◽  
pp. 358-366 ◽  
Author(s):  
Huriye Erdogan
Keyword(s):  
Cytochrome P450 ◽  
Time Course ◽  
Catalytic Cycle ◽  
Small Step ◽  
Compound I ◽  
Unique Role ◽  
Natural Time ◽  
Giant Leap ◽  
High Valent

The intermediates operating in the cytochrome P450 catalytic cycle have been investigated for more than half a century, fascinating many enzymologists. Each intermediate has its unique role to carry out diverse oxidations. Natural time course of the catalytic cycle is quite fast, hence, not all of the reactive intermediates could be isolated during physiological catalysis. Different high-valent iron intermediates have been proposed as primary oxidants: the candidates are compound 0 (Cpd 0, [FeOOH][Formula: see text]P450) and compound I (Cpd I, Fe(IV)[Formula: see text]O por[Formula: see text]P450). Among them, the role of Cpd I in hydroxylation is fairly well understood due the discovery of the peroxide shunt. This review endeavors to put the outstanding research efforts conducted to isolate and characterize the intermediates together. In addition to spectral features of each intermediate in the catalytic cycle, the oxidizing powers of Cpd 0 and Cpd I will be discussed along with most recent scientific findings.

scholarly journals Cryotrapped Reaction Intermediates of Cytochrome P450 Studied by Radiolytic Reduction with Phosphorus-32

2001 ◽  
Vol 276 (15) ◽  
pp. 11648-11652 ◽  
Author(s):  
Ilia G. Denisov ◽  
Thomas M. Makris ◽  
Stephen G. Sligar
Keyword(s):  
Cytochrome P450 ◽  
Catalytic Cycle ◽  
Internal Source ◽  
Reaction Intermediates ◽  
Compound I ◽  
Γ Irradiation ◽  
Rate Limiting Step ◽  
Type Spectrum ◽  
Radiolytic Reduction ◽  
Electron Reduction

Unstable reaction intermediates of the cytochrome P450 catalytic cycle have been prepared at cryogenic temperatures using radiolytic one-electron reduction of the oxy-P450 CYP101 complex. Since a rate-limiting step in the catalytic cycle of the enzyme is the reduction of the ferrous oxygenated heme protein, subsequent reaction intermediates do not normally accumulate. Using60Co γ-irradiation, the primary reduced oxy-P450 species at 77 K has been identified as a superoxo- or hydroperoxo-Fe3+-heme complex (Davydov, R., Macdonald, I. D. G., Makris, T. M., Sligar, S. G., and Hoffman, B. M. (1999)J. Am. Chem. Soc.121, 10654–10655). The electronic absorption spectroscopy is an essential tool to characterize cytochrome P450 intermediates and complements paramagnetic methods, which are blind to important diamagnetic or antiferromagnetically coupled states. We report a method of trapping unstable states of redox enzymes using phosphorus-32 as an internal source of electrons. We determine the UV-visible optical spectra of the reduced oxygenated state of CYP101 and show that the primary intermediate, a hydroperoxo-P450, is stable below 180 K and converts smoothly to the product complex at ∼195 K. In the course of the thermal annealing, no spectral changes indicating the presence of oxoferryl species (the so-called compound I type spectrum) was observed.

scholarly journals Redox-Linked Domain Movements in the Catalytic Cycle of Cytochrome P450 Reductase

Structure ◽  
2013 ◽  
Vol 21 (9) ◽  
pp. 1581-1589 ◽  
Author(s):  
Wei-Cheng Huang ◽  
Jacqueline Ellis ◽  
Peter C.E. Moody ◽  
Emma L. Raven ◽  
Gordon C.K. Roberts
Keyword(s):  
Cytochrome P450 ◽  
Catalytic Cycle ◽  
Cytochrome P450 Reductase ◽  
P450 Reductase

scholarly journals A novel approach to distinguish between enzyme mechanisms: quasi-steady-state kinetic analysis of the prostaglandin H synthase peroxidase reaction

2003 ◽  
Vol 372 (3) ◽  
pp. 713-724 ◽  
Author(s):  
Peter V. VRZHESHCH ◽  
Elena A. BATANOVA ◽  
Alevtina T. MEVKH ◽  
Sergei D. VARFOLOMEEV ◽  
Irina G. GAZARYAN ◽  
...  
Keyword(s):  
Experimental Data ◽  
Steady State ◽  
Catalytic Cycle ◽  
Prostaglandin H Synthase ◽  
Compound I ◽  
Novel Approach ◽  
Steady State Kinetic ◽  
Enzyme Intermediates ◽  
Prostaglandin H ◽  
State Kinetic

A method of analysis for steady-state kinetic data has been developed that allows relationships between key partial reactions in the catalytic cycle of a functioning enzyme to be determined. The novel approach is based on a concept of scalar and vector ‘kinetic connectivities’ between enzyme intermediates in an arbitrary enzyme mechanism. The criterion for the agreement between experimental data and a proposed kinetic model is formulated as the kinetic connectivity of intermediate forms of the enzyme. This concept has advantages over conventional approaches and is better able to describe the complex kinetic behaviour of prostaglandin H synthase (PGHS) when catalysing the oxidation of adrenaline by H2O2. To interpret the experimental data for PGHS, a generalized model for multi-substrate enzyme reactions was developed with provision for irreversible enzyme inactivation. This model showed that two enzyme intermediates must undergo inactivation during the catalytic cycle. These forms are proposed to be PGHS compound I and a compound I–adrenaline complex.

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