Resonance Raman Spectroscopy of the Oxygenated Intermediates of Human CYP19A1 Implicates a Compound I Intermediate in the Final Lyase Step

Mak, Piotr J.; Luthra, Abhinav; Sligar, Stephen G.; Kincaid, James R.

doi:10.1021/ja500054c

Cited by 51 publications

(74 citation statements)

References 36 publications

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“…Furthermore, a very recent resonance Raman study found no significant difference between the oxy-complexes of aromatase in the first and third sub-cycles implicating the involvement of Compound I in the terminal lyase step. 27 Kinetic solvent isotope effect studies of the three sub-cycles of aromatase led to an identical conclusion. 28 Both experimental and theoretical studies clearly indicate that the reactivity of CYPs is not a clear-cut mechanism.…”

Section: 19mentioning

confidence: 81%

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The mechanism of human aromatase (CYP 19A1) revisited: DFT and QM/MM calculations support a compound I-mediated pathway for the aromatization process

Krámos

Oláh

2014

Struct Chem

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Human aromatase is responsible for the last step of estrogen biosynthesis, for the aromatization of ring A of androstenedione (ASD) or testosterone. In this work the mechanism of aromatization was studied using gas phase and hybrid QM/MM calculations. It is shown that human aromatase can efficiently catalyze the aromatization process via a Compound I (or Compound II) mediated pathway. The nature of the oxidant is very sensitive to the polarizing environment of the enzyme, as the oxidant has a Compound I nature in the gas phase * corresponding author, to whom corresponding should be addressed e-mail: julianna.olah@mail.bme.hu; telephone number: +36-1-463-1286 2 calculations, which is modulated by the enzyme environment to become a mixed Compound I and Compound II character. The electronic structure of the obtained QM-only and QM/MM stationary points is thoroughly discussed.

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Section: 19mentioning

confidence: 81%

“…27,28 A significant difference has been found in the reaction mechanism as predicted by QM-only and QM/MM calculations. According to the QM-only calculations 1β-hydrogen is abstracted by …”

Section: Resultsmentioning

confidence: 99%

The mechanism of human aromatase (CYP 19A1) revisited: DFT and QM/MM calculations support a compound I-mediated pathway for the aromatization process

Krámos

Oláh

2014

Struct Chem

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“…An example is the use of resonance Raman spectroscopy to reveal control of regio- and stereo- specificity of catalysis, which would be impossible with a highly heterogeneous or aggregated sample. 82,86,87,89 …”

Section: Application Of Nanodiscs In Functional Studies Of Membranmentioning

confidence: 99%

Nanodiscs in Membrane Biochemistry and Biophysics

2017

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Membrane proteins play a most important part in metabolism, signaling, cell motility, transport, development, and many other biochemical and biophysical processes which constitute fundamentals of life on molecular level. Detailed understanding of these processes is necessary for the progress of life sciences and biomedical applications. Nanodiscs provide a new and powerful tool for a broad spectrum of biochemical and biophysical studies of membrane proteins and are commonly acknowledged as an optimal membrane mimetic system which provides control over size, composition and specific functional modifications on nanometer scale. In this review we attempted to combine a comprehensive list of various applications of Nanodisc technology with systematic analysis of the most attractive features of this system and advantages provided by Nanodiscs for structural and mechanistic studies of membrane proteins.

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“…An exhaustive list of spectroscopic techniques will not be discussed in this section, but another technique worth mentioning is resonance Raman spectroscopy, which selectively measures the vibrational modes of the heme group in P450 enzymes. This technique was used to observe the different P450 species depending on the substrate used [25, 26]. …”

Section: Stopped-flow Technique (Steps 1 2 9 – Substrate Bindingmentioning

confidence: 99%

Rapid kinetic methods to dissect steroidogenic cytochrome P450 reaction mechanisms

Yoshimoto

Auchus

2016

The Journal of Steroid Biochemistry and Molecular Biology

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All cytochrome P450 enzyme reactions involve a catalytic cycle with several discreet physical or chemical steps. This cycle ends with the formation of the reactive heme iron-oxygen complex, which oxygenates substrate. While the steps might be very similar for each P450 enzyme, the rates of each step varies tremendously for each enzyme and sometimes even for different reactions catalyzed by the same enzyme. For example, the rate-limiting step for most bacterial P450 enzymes, with turnover numbers over 1,000 sec−1, is the second electron transfer. In contrast, steroidogenic P450s from eukaryotes catalyze much slower reactions, with turnover numbers of ~5–250 min−1; therefore, assumptions about kinetic properties for the mammalian P450 enzymes based on the bacterial enzymes are tenuous. In order to dissect the rates for individual steps, special techniques that isolate individual steps and/or single turnovers are required. This article will review the theoretical principles and practical considerations for several of these techniques, with illustrative published examples. The reader should gain an appreciation for the appropriate methods used to interrogate particular steps in the P450 reaction cycle.

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Resonance Raman Spectroscopy of the Oxygenated Intermediates of Human CYP19A1 Implicates a Compound I Intermediate in the Final Lyase Step

Cited by 51 publications

References 36 publications

The mechanism of human aromatase (CYP 19A1) revisited: DFT and QM/MM calculations support a compound I-mediated pathway for the aromatization process

The mechanism of human aromatase (CYP 19A1) revisited: DFT and QM/MM calculations support a compound I-mediated pathway for the aromatization process

Nanodiscs in Membrane Biochemistry and Biophysics

Rapid kinetic methods to dissect steroidogenic cytochrome P450 reaction mechanisms

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