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Abad, José Luís; Camps, Francisco; Fabriàs, Gemma

doi:10.1002/1521-3757(20000915)112:18<3417::aid-ange3417>3.3.co;2-d

Cited by 6 publications

(7 citation statements)

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“…The other approach involves a direct C−H bond activation process where metal cofactors, such as heme‐iron centers, non‐heme mono‐iron centers, or di‐iron centers, are required to trigger the reaction . For two mechanistically characterized desaturases, fatty acyl desaturases and cytochrome P450 enzymes (Scheme B), enzymatic and biomimetic model studies support a mechanism involving two consecutive hydrogen atom transfer processes (2‐HAT) to construct a C=C bond –,,. Specifically, a di‐(μ‐oxo)‐di‐iron(IV/IV) intermediate or a ferryl porphyrin radical cation species, compound I, functions as the key intermediate to trigger the first C−H bond cleavage, and a proposed Fe 2 III/IV intermediate or an Fe IV ‐hydroxo complex, compound II, is hypothesized to serve as the second hydrogen atom (H ⋅ ) abstractor to produce a di‐radical species prior to C=C bond formation.…”

Section: Methodsmentioning

confidence: 98%

Insights into the Desaturation of Cyclopeptin and its C3 Epimer Catalyzed by a non‐Heme Iron Enzyme: Structural Characterization and Mechanism Elucidation

Liao

Huang

et al. 2018

Angew Chem Int Ed

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AsqJ, an iron(II)- and 2-oxoglutarate-dependent enzyme found in viridicatin-type alkaloid biosynthetic pathways, catalyzes sequential desaturation and epoxidation to produce cyclopenins. Crystal structures of AsqJ bound to cyclopeptin and its C3 epimer are reported. Meanwhile, a detailed mechanistic study was carried out to decipher the desaturation mechanism. These findings suggest that a pathway involving hydrogen atom abstraction at the C10 position of the substrate by a short-lived Fe -oxo species and the subsequent formation of a carbocation or a hydroxylated intermediate is preferred during AsqJ-catalyzed desaturation.

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Section: Methodsmentioning

confidence: 98%

Insights into the Desaturation of Cyclopeptin and its C3 Epimer Catalyzed by a non‐Heme Iron Enzyme: Structural Characterization and Mechanism Elucidation

Liao

Huang

et al. 2018

Angew Chem Int Ed

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“…Buried water molecules have been assigned to important roles in biological function, including ligand binding, protein stability and flexibility. [53][54][55][56][57] In the case of buried waters that become displaced upon ligand binding, 58 these can provide an important thermodynamic driving force for enhanced substrate affinity (see for examples, [55][56][57]59 ). WT SLO has at least five X-ray-resolved water molecules that line the putative ligand binding pocket 29 and most, if not all, are likely to be expelled upon substrate acquisition.…”

Section: Origins Of the Elevated Enthalpic Barriers In Slo Mutantsmentioning

confidence: 99%

Biophysical Characterization of a Disabled Double Mutant of Soybean Lipoxygenase: The “Undoing” of Precise Substrate Positioning Relative to Metal Cofactor and an Identified Dynamical Network

et al. 2019

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Soybean lipoxygenase (SLO) has served as a prototype for understanding the molecular origin of enzymatic rate accelerations. The double mutant (DM) L546A/L754A is considered a dramatic outlier, due to the unprecedented size and near temperature-independence of its primary kinetic isotope effect, low catalytic efficiency, and elevated enthalpy of activation. To uncover the physical basis of these features, we herein apply three structural probes: hydrogen-deuterium exchange mass spectrometry, room-temperature X-ray crystallography and EPR spectroscopy on four SLO variants (wild-type (WT) enzyme, DM, and the two parental single mutants, L546A and L754A). DM is found to incorporate features of each parent, with the perturbation at position 546 predominantly influencing thermally activated motions that connect the active site to a proteinsolvent interface, while mutation at position 754 disrupts the ligand field and solvation near the cofactor iron. However, the expanded active site in DM leads to more active site water molecules *

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“…It is now widely accepted that H tunneling (proton, hydrogen, or hydride) occurs in enzyme‐catalyzed reactions,1, 6–11 but the role of promoting motions in modulating the tunneling barrier remains contentious 3. Experimental identification of coupled (promoting) motions is challenging, with experimental evidence for environmentally coupled H‐tunneling reactions mainly inferred from the unusual temperature dependence of primary kinetic isotope effects (KIEs) 12–15. In addition to temperature,12, 13 there are other intensive (bulk) properties that, in principle, can be used to probe these reactions, including pressure10, 16 and solution viscosity 17–21.…”

Section: The Effect Of Solution Viscosity On the Saturation Kinetics mentioning

confidence: 99%

Are Environmentally Coupled Enzymatic Hydrogen Tunneling Reactions Influenced by Changes in Solution Viscosity?

et al. 2007

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The role and importance of protein dynamics in enzymatic reactions remains a key question in enzymology. [1][2][3][4][5] Of great current interest is whether enzymes have evolved to use quantum tunneling to the best advantage by, when necessary, coupling specific protein motions (vibrations) to the tunneling reaction coordinate. It is now widely accepted that H tunneling (proton, hydrogen, or hydride) occurs in enzymecatalyzed reactions, [1,[6][7][8][9][10][11] but the role of promoting motions in modulating the tunneling barrier remains contentious. [3] Experimental identification of coupled (promoting) motions is challenging, with experimental evidence for environmentally coupled H-tunneling reactions mainly inferred from the unusual temperature dependence of primary kinetic isotope effects (KIEs). [12][13][14][15] In addition to temperature, [12,13] there are other intensive (bulk) properties that, in principle, can be used to probe these reactions, including pressure [10,16] and solution viscosity. [17][18][19][20][21] We have demonstrated the utility of combining pressure and temperature to study environmentally coupled H tunneling in the reductive half-reaction (RHR) of morphinone reductase (MR), involving hydride tunneling from the coenzyme NADH to the enzyme-bound cofactor flavin mononucleotide (FMN).[10] Although changes in solution viscosity have been used to probe protein rearrangement after CO dissociation from myoglobin, [21] configurational and conformational gating of interprotein electron-transfer (ET) reactions, [17][18][19][20] and conformational gating of intraprotein ET, [22] studies of the dynamical influence of solvent viscosity on enzymatic H-tunneling reactions are less common. Moreover, there has been no rigorous quantitative analysis of these effects.At a phenomenological level, we have shown that the magnitude and temperature dependence of the reaction rate and KIE measured for proton tunneling in the RHR of methylamine dehydrogenase (MADH) are unchanged following the addition of 30 % glycerol (viscosity increase ca. 2-3-fold) to the reaction solution.[12] Similarly, a decrease in KIE and increase in apparent enthalpy for the RHR of l-phenylalanine oxidase (PAO) upon the addition of 30 % glycerol has been reported.[23] Glycosylation of a protein surface can also affect the dynamic motion of a protein, and this approach has been used to study the rate of hydride transfer in glucose oxidase (GO) [24,25] by 1) using various glycoforms of the enzyme that differ in the extent of glycosylation [24] and 2) replacing the native polysaccharide with different polymeric forms of polyethylene glycol.[25] By both increasing and decreasing the apparent surface viscosity, decreases in the fitness of the enzyme were observed, reducing the Arrhenius pre-exponential ratio (A D /A T ) for deuterium and tritium transfer away from unity. [24,25] GO, MADH, and PAO do not have strongly temperature-dependent KIEs, suggesting a minor role (if any) for fast, non-equilibrated promoting motions coupled to the H-tr...

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Cited by 6 publications

References 0 publications

Insights into the Desaturation of Cyclopeptin and its C3 Epimer Catalyzed by a non‐Heme Iron Enzyme: Structural Characterization and Mechanism Elucidation

Insights into the Desaturation of Cyclopeptin and its C3 Epimer Catalyzed by a non‐Heme Iron Enzyme: Structural Characterization and Mechanism Elucidation

Biophysical Characterization of a Disabled Double Mutant of Soybean Lipoxygenase: The “Undoing” of Precise Substrate Positioning Relative to Metal Cofactor and an Identified Dynamical Network

Are Environmentally Coupled Enzymatic Hydrogen Tunneling Reactions Influenced by Changes in Solution Viscosity?

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