Adriana Dinescu

Copper–Nitrene Complexes in Catalytic CH Amination†

The direct catalytic transformation of typically inert carbon– hydrogen bonds into carbon–oxygen, carbon–nitrogen, and carbon–carbon bonds represents a significant challenge in organic synthesis and the chemical industry. Ideally, such reactions would not require the presence of a functional group that is discarded later in the course of the chemical transformation, but would proceed with atom and energy efficiency and be of minimal environmental impact. Chemical species capable of directly promoting this transformation often contain multiple bonds between late transition metals to oxygen, nitrogen, or carbon centers. Nature provides inspiration with enzymes such as cyctochrome P-450, which is efficient at inserting an oxygen atom into hydrocarbon C H bonds through an iron–oxo intermediate ([Fe]=O). The use of iminoiodanes (PhI=NR) has led to the generation of related metal–nitrene intermediates ([M]=NR), which in certain cases, exhibit stoichiometric reactivity with carbon–hydrogen bonds to give amines. Several catalytic protocols based on Rh, Ru, Ag, Au, Cu, and other metals for the amination of benzylic or allylic C H bonds with iminoiodanes bearing electrondeficient N substituents represent useful methodologies for the transformation of C H into C N bonds. 10,11] In these catalytic cases, metal–nitrene species are inferred, but their high reactivity makes their characterization as the active intermediates difficult. This lack of characterization is unfortunate, as a fundamental understanding of metal–nitrene species that mediate catalytic C N bond formation would aid the development of more general systems for C H bond functionalization. In this context, we recently isolated a dicopper complex [{(Me3NN)Cu}2(m-NAr)] (Ar = 3,5-dimethylphenyl) in which a nitrene is bound between two b-diketiminate copper fragments. Solution studies supported the intermediacy of terminal copper–nitrene [Cu] = [(Me3NN)Cu] through the dissociation of a b-diketiminate copper fragment from [{(Me3NN)Cu}2(m-NAr)]. The terminal nitrene could not be directly observed owing to an unfavorable dissociation constant [Eq. (1); [Cu] = [(Me3NN)Cu].

The correlation-consistent composite approach: Application to the G3/99 test set

The correlation-consistent composite approach (ccCA), an ab initio composite technique for computing atomic and molecular energies, recently has been shown to successfully reproduce experimental data for a number of systems. The ccCA is applied to the G3/99 test set, which includes 223 enthalpies of formation, 88 adiabatic ionization potentials, 58 adiabatic electron affinities, and 8 adiabatic proton affinities. Improvements on the original ccCA formalism include replacing the small basis set quadratic configuration interaction computation with a coupled cluster computation, employing a correction for scalar relativistic effects, utilizing the tight-d forms of the second-row correlation-consistent basis sets, and revisiting the basis set chosen for geometry optimization. With two types of complete basis set extrapolation of MP2 energies, ccCA results in an almost zero mean deviation for the G3/99 set (with a best value of -0.10 kcal mol(-1)), and a 0.96 kcal mol(-1) mean absolute deviation, which is equivalent to the accuracy of the G3X model chemistry. There are no optimized or empirical parameters included in the computation of ccCA energies. Except for a few systems to be discussed, ccCA performs as well as or better than Gn methods for most systems containing first-row atoms, while for systems containing second-row atoms, ccCA is an improvement over Gn model chemistries.

Bonding and Structure of Copper Nitrenes

Copper nitrenes are of interest as intermediates in the catalytic aziridination of olefins and the amination of C-H bonds. However, despite advances in the isolation and study of late-transition-metal multiply bonded complexes, a bona fide structurally characterized example of a terminal copper nitrene has, to our knowledge, not been reported. In anticipation of such a report, terminal copper nitrenes are studied from a computational perspective. The nitrene complexes studied here are of the form (beta-diketiminate)Cu(NPh). Density functional theory (DFT), complete active space self-consistent-field (CASSCF) electronic structure techniques, and hybrid quantum mechanical/molecular mechanical (QM/MM) methods are employed to study such species. While DFT methods indicate that a triplet (S = 1) is the ground state, CASSCF calculations indicate that a singlet (S = 0) is the ground state, with only a small energy gap between the singlet and triplet. Moreover, the ground-state (open-shell) singlet copper nitrene is found to be highly multiconfigurational (i.e., biradical) and to possess a bent geometry about the nitrene nitrogen, contrasting with the linear nitrene geometry of the triplet copper nitrenes. CASSCF calculations also reveal the existence of a closed-shell singlet state with some degree of multiple bonding character for the copper-nitrene bond.

Thermodynamic and Structural Features of Aqueous Ce(III)

With a single f-electron, Ce(III) is the simplest test case for benchmarking the thermodynamic and structural properties of hydrated Ln(III) against varying density functionals and reaction field models, in addition to determining the importance of multiconfigurational character in their wave functions. Here, the electronic structure of Ce(H2O)x(H 2O)y(3+) (x = 8, 9; y = 0, 12-14) has been examined using DFT and CASSCF calculations. The latter confirmed that the wave function of octa- and nona-aqua Ce(III) is well-described by a single configuration. Benchmarking was performed for density functionals, reaction field cavity types, and solvation reactions against the experimental free energy of hydration, DeltaG(hyd)(Ce(3+)). The UA0, UAKS, Pauling, and UFF polarized continuum model cavities displayed different performance, depending on whether one or two hydration shells were examined, and as a function of the size of the metal basis set. These results were essentially independent of the density functional employed. Using these benchmarks, the free energy for water exchange between CN = 8 and CN = 9, for which no experimental data are available, was estimated to be approximately -4 kcal/mol.

Function of Conserved Residues of Human Glutathione Synthetase IMPLICATIONS FOR THE ATP-grasp ENZYMES

Glutathione synthetase is an enzyme that belongs to the glutathione synthetase ATP-binding domain-like superfamily. It catalyzes the second step in the biosynthesis of glutathione from γ-glutamylcysteine and glycine in an ATP-dependent manner. Glutathione synthetase has been purified and sequenced from a variety of biological sources; still, its exact mechanism is not fully understood. A variety of structural alignment methods were applied and four highly conserved residues of human glutathione synthetase (Glu-144, Asn-146, Lys-305, and Lys-364) were identified in the binding site. The function of these was studied by experimental and computational site-directed mutagenesis. The three-dimensional coordinates for several human glutathione synthetase mutant enzymes were obtained using molecular mechanics and molecular dynamics simulation techniques, starting from the reported crystal structure of human glutathione synthetase. Consistent with circular dichroism spectroscopy, our results showed no major changes to overall enzyme structure upon residue mutation. However, semiempirical calculations revealed that ligand binding is affected by these mutations. The key interactions between conserved residues and ligands were detected and found to be essential for enzymatic activity. Particularly, the negatively charged Glu-144 residue plays a major role in catalysis.

Determination of the active site of Sphingobium chlorophenolicum 2,6-dichlorohydroquinone dioxygenase (PcpA)

Abstract2,6-Dichlorohydroquinone 1,2-dioxygenase (PcpA) from Sphingobium chlorophenolicum ATCC 39723 is a member of a class of Fe(II)-containing hydroquinone dioxygenases that is involved in the mineralization of the pollutant pentachlorophenol. This enzyme has not been extensively characterized, despite its interesting ring-cleaving activity and use of Fe(II), which are reminiscent of the well-known extradiol catechol dioxygenases. On the basis of limited sequence homology to the extradiol catechol dioxygenases, the residues ligating the Fe(II) center were originally proposed to be H159, H227, and E276 (Xu et al. in Biochemistry 38:7659–7669, 1999). However, PcpA has higher sequence homology to a newly reported, crystallographically characterized zinc metalloenzyme that has a similar predicted fold. We generated a homology model of the structure of PcpA based upon the structure of this zinc metalloenzyme. The homology model predicts that the tertiary structure of PcpA differs significantly from that of the extradiol dioxygenases, and that the residues ligating the Fe(II) are H11, H227, and E276. This structural model was tested by mutating each of H11, H159, H227, and E276 to alanine. An additional residue that is predicted to lie near the active site and is conserved among PcpA, its closest homologues, and the extradiol dioxygenases, Y266, was mutated to phenylalanine. Of these mutants, only H159A retained significant activity, thus confirming the active-site location predicted by the homology-based structural model. The model provides an important basis for understanding the origin of the unique function of PcpA.

Investigation of Uranyl Nitrate Ion Pairs Complexed with Amide Ligands using Electrospray Ionization Ion Trap Mass Spectrometry and Density Functional Theory

Ion populations formed from electrospray of uranyl nitrate solutions containing different amides vary depending on ligand nucleophilicity and steric crowding at the metal center. The most abundant species were ion pair complexes having the general formula [UO(2)(NO(3))(amide)(n=2,3)](+); however, singly charged complexes containing the amide conjugate base and reduced uranyl UO(2)(+) were also formed as were several doubly charged species. The formamide experiment produced the greatest diversity of species resulting from weaker amide binding, leading to dissociation and subsequent solvent coordination or metal reduction. Experiments using methyl formamide, dimethyl formamide, acetamide, and methyl acetamide produced ion pair and doubly charged complexes that were more abundant and less abundant complexes containing solvent or reduced uranyl. This pattern is reversed in the dimethylacetamide experiment, which displayed lower abundance doubly charged complexes, but augmented reduced uranyl complexes. DFT investigations of the tris-amide ion pair complexes showed that interligand repulsion distorts the amide ligands out of the uranyl equatorial plane and that complex stabilities do not increase with increasing amide nucleophilicity. Elimination of an amide ligand largely relieves the interligand repulsion, and the remaining amide ligands become closely aligned with the equatorial plane in the structures of the bis-amide ligands. The studies show that the phenomenological distribution of coordination complexes in a metal-ligand electrospray experiment is a function of both ligand nucleophilicity and interligand repulsion and that the latter factor begins exerting influence even in the case of relatively small ligands like the substituted methyl-formamide and methyl-acetamide ligands.

The Role of the Glycine Triad in Human Glutathione Synthetase

Experimental kinetics and computational modeling of human glutathione synthetase (hGS) support the significant role of the G-loop glycine triad (G369, G370, G371) for activity of this ATP-grasp enzyme. Enzyme kinetic experiments indicate that G369V and G370V mutant hGS have little activity (<0.7 and 0.3%, respectively, versus wild-type hGS). However, G371V retains ∼13% of the activity of wild-type hGS. With respect to G-loop:A-loop interaction in hGS, mutations at Gly369 and Gly370 decrease ligand binding and prevent active site closure and protection. This research indicates that Gly369 and Gly370 have essential roles in hGS, while Gly371 has a lesser involvement. Implications for glycine-rich ensembles in other phosphate-binding enzymes are discussed.

ASPARTATE 458 of Human Glutathione Synthetase is Important for Cooperativity and Active Site Structure

Human glutathione synthetase (hGS) catalyzes the second ATP-dependent step in the biosynthesis of glutathione (GSH) and is negatively cooperative to the γ-glutamyl substrate. The hGS active site is composed of three highly conserved catalytic loops, notably the alanine rich A-loop. Experimental and computational investigations of the impact of mutation of Asp458 are reported, and thus the role of this A-loop residue on hGS structure, activity, negativity cooperativity and stability is defined. Several Asp458 hGS mutants (D458A, D458N and D458R) were constructed using site-directed mutagenesis and their activities determined (10%, 15% and 7% of wild-type hGS, respectively). The Michaelis-Menten constant (K(m)) was determined for all three substrates (glycine, GAB and ATP): glycine K(m) increased by 30-115-fold, GAB K(m) decreased by 8-17-fold, and the ATP K(m) was unchanged. All Asp458 mutants display a change in cooperativity from negative cooperativity to non-cooperative. All mutants show similar stability as compared to wild-type hGS, as determined by differential scanning calorimetry. The findings indicate that Asp458 is essential for hGS catalysis and that it impacts the allostery of hGS.

Electronic Structure Studies on Deprotonation of Dithiophosphinic Acids in Water Clusters

We report herein a computational study of proton transfer reactions between dithiophosphinic acids (HAs) and water clusters using B3LYP and MP2 methods. The ground-state and transition-state structures of HA-(H(2)O)(n) (n = 1, 2, 3) cluster complexes have been calculated. The influence of water molecules on energy barrier heights of proton transfer reactions has been examined in the gas phase and solution for bis[o-(trifluoromethyl)phenyl]- and bis(2,4,4-trimethylpentyl)dithiophosphinic acids (HA1 and HA2, respectively). Gas-phase calculations indicate that electron-withdrawing substituents and trifluoromethyl groups in the ortho position favor deprotonation of HA1 when three water molecules are included in the cluster. This suggests that at least three water molecules are necessary to solvate the abstracted proton in the presence of the anion. In the case of HA2, the electron-donating groups favor the reverse proton transfer reaction, namely, protonation of dithiophosphinate anion. Bulk solvent effects have been modeled for aqueous and organic media with the CPCM model. The calculated results show that polar solvents can lower the activation energy for less energetically stable transition states that have more localized charges.