Systematically testing human HMBS missense variants to reveal mechanism and pathogenic variation.

Defects in hydroxymethylbilane synthase (HMBS) can cause acute intermittent porphyria (AIP), an acute neurological disease. Although sequencing-based diagnosis can be definitive, ∼⅓ of clinical HMBS variants are missense variants, and most clinically reported HMBS missense variants are designated as "variants of uncertain significance" (VUSs). Using saturation mutagenesis, en masse selection, and sequencing, we applied a multiplexed validated assay to both the erythroid-specific and ubiquitous isoforms of HMBS, obtaining confident functional impact scores for >84% of all possible amino acid substitutions. The resulting variant effect maps generally agreed with biochemical expectations and provide further evidence that HMBS can function as a monomer. Additionally, the maps implicated specific residues as having roles in active site dynamics, which was further supported by molecular dynamics simulations. Most importantly, these maps can help discriminate pathogenic from benign HMBS variants, proactively providing evidence even for yet-to-be-observed clinical missense variants.

Identification of Catechins' Binding Sites in Monomeric Aß42 through Ensemble Docking and MD Simulations.

The assembly of the amyloid-ß peptide (Aß) into toxic oligomers and fibrils is associated with Alzheimer's disease and dementia. Therefore, disrupting amyloid assembly by direct targeting of the Aß monomeric form with small molecules or antibodies is a promising therapeutic strategy. However, given the dynamic nature of Aß, standard computational tools cannot be easily applied for high-throughput structure-based virtual screening in drug discovery projects. In the current study, we propose a computational pipeline-in the framework of the ensemble docking strategy-to identify catechins' binding sites in monomeric Aß42. It is shown that both hydrophobic aromatic interactions and hydrogen bonding are crucial for the binding of catechins to Aß42. Additionally, it has been found that all the studied ligands, especially EGCG, can act as potent inhibitors against amyloid aggregation by blocking the central hydrophobic region of Aß. Our findings are evaluated and confirmed with multi-microsecond MD simulations. Finally, it is suggested that our proposed pipeline, with low computational cost in comparison with MD simulations, is a suitable approach for the virtual screening of ligand libraries against Aß.

Systematically testing human HMBS missense variants to reveal mechanism and pathogenic variation.

Defects in hydroxymethylbilane synthase (HMBS) can cause Acute Intermittent Porphyria (AIP), an acute neurological disease. Although sequencing-based diagnosis can be definitive, ~⅓ of clinical HMBS variants are missense variants, and most clinically-reported HMBS missense variants are designated as "variants of uncertain significance" (VUS). Using saturation mutagenesis, en masse selection, and sequencing, we applied a multiplexed validated assay to both the erythroid-specific and ubiquitous isoforms of HMBS, obtaining confident functional impact scores for >84% of all possible amino-acid substitutions. The resulting variant effect maps generally agreed with biochemical expectation. However, the maps showed variants at the dimerization interface to be unexpectedly well tolerated, and suggested residue roles in active site dynamics that were supported by molecular dynamics simulations. Most importantly, these HMBS variant effect maps can help discriminate pathogenic from benign variants, proactively providing evidence even for yet-to-be-observed clinical missense variants.

Structural Investigation of DHICA Eumelanin Using Density Functional Theory and Classical Molecular Dynamics Simulations.

Eumelanin is an important pigment, for example, in skin, hair, eyes, and the inner ear. It is a highly heterogeneous polymer with 5,6-dihydroxyindole-2-carboxylic acid (DHICA) and 5,6-dihydroxyindole (DHI) building blocks, of which DHICA is reported as the more abundant in natural eumelanin. The DHICA-eumelanin protomolecule consists of three building blocks, indole-2-carboxylic acid-5,6-quinone (ICAQ), DHICA and pyrrole-2,3,5-tricarboxylic acid (PTCA). Here, we focus on the self-assembly of DHICA-eumelanin using multi-microsecond molecular dynamics (MD) simulations at various concentrations in aqueous solutions. The molecule was first parameterized using density functional theory (DFT) calculations. Three types of systems were studied: (1) uncharged DHICA-eumelanin, (2) charged DHICA-eumelanin corresponding to physiological pH, and (3) a binary mixture of both of the above protomolecules. In the case of uncharged DHICA-eumelanin, spontaneous aggregation occurred and water molecules were present inside the aggregates. In the systems corresponding to physiological pH, all the carboxyl groups are negatively charged and the DHICA-eumelanin model has a net charge of -4. The effect of K+ ions as counterions was investigated. The results show high probability of binding to the deprotonated oxygens of the carboxylate anions in the PTCA moiety. Furthermore, the K+ counterions increased the solubility of DHICA-eumelanin in its charged form. A possible explanation is that the charged protomolecules favor binding to the K+ ions rather than aggregating and binding to other protomolecules. The binary mixtures show aggregation of uncharged DHICA-eumelanins; unlike the charged systems with no aggregation, a few charged DHICA-eumelanins are present on the surface of the uncharged aggregation, binding to the K+ ions.

Collective interactions among organometallics are exotic bonds hidden on lab shelves.

Recent discovery of an unusual bond between Na and B in NaBH3- motivated us to look for potentially similar bonds, which remained unnoticed among systems isoelectronic with NaBH3-. Here, we report a novel family of collective interactions and a measure called exchange-correlation interaction collectivity index (ICIXC; [Formula: see text]) to characterize the extent of collective versus pairwise bonding. Unlike conventional bonds in which ICIXC remains close to one, in collective interactions ICIXC may approach zero. We show that collective interactions are commonplace among widely used organometallics, as well as among boron and aluminum complexes with the general formula [Ma+AR3]b- (A: C, B or Al). In these species, the metal atom interacts more efficiently with the substituents (R) on the central atoms than the central atoms (A) upon forming efficient collective interactions. Furthermore, collective interactions were also found among fluorine atoms of XFn systems (X: B or C). Some of organolithium and organomagnesium species have the lowest ICIXC among the more than 100 studied systems revealing the fact that collective interactions are rather a rule than an exception among organometallic species.

Free Energy and Stacking of Eumelanin Nanoaggregates.

Eumelanin, a member of the melanin family, is a black-brown insoluble pigment. It possesses a broad range of properties such as antioxidation, free radical scavenging, photoprotection, and charge carrier transportation. Surprisingly, the exact molecular structure of eumelanin remains undefined. It is, however, generally considered to consist of two main building blocks, 5,6-dihydroxyindole (DHI) and 5,6- dihydroxyindole carboxylic acid (DHICA). We focus on DHI and report, for the first time, a computational investigation of the structural properties of DHI-eumelanin aggregates in aqueous solutions. First, multimicrosecond molecular dynamics (MD) simulations at different concentrations were performed to investigate the aggregation and ordering of tetrameric DHI-eumelanin protomolecules. This was followed by umbrella sampling (US) and density functional theory (DFT) calculations to study the physical mechanisms of stacking. Aggregation occurs through formation of nanoscale stacks and was observed in all systems. Further analyses showed that aggregation and coarsening of the domains is due to a decrease in hydrogen bonds between the eumelanins and water; while domains exist, there is no long-range order. The results show noncovalent stacks with the interlayer distance between eumelanin protomolecules being less than 3.5 Å. This is in good agreement with transmission electron microscopy data. Both free energy calculations and DFT revealed strong stacking interactions. The electrostatic potential map provides an explanation and a rationale for the slightly sheared relative orientations and, consequently, for the curved shapes of the nanoscale domains.

Dawn of a New Era for Membrane Protein Design.

A major advancement has recently occurred in the ability to predict protein secondary structure from sequence using artificial neural networks. This new accessibility to high-quality predicted structures provides a big opportunity for the protein design community. It is particularly welcome for membrane protein design, where the scarcity of solved structures has been a major limitation of the field for decades. Here, we review the work done to date on the membrane protein design and set out established and emerging tools that can be used to most effectively exploit this new access to structures.

Biphasic Proton Transport Mechanism for Uncoupling Proteins.

It has been suggested that uncoupling proteins (UCPs) transport protons via interconversion between two conformational states: one in the "cytoplasmic state" and the other in the "matrix state". Matrix and cytoplasmic salt-bridge networks are key controllers of these states. This study proposes a mechanism for proton transport in tetrameric UCP2, with focus on the role of the matrix network. Eleven mutants were prepared to disrupt (K → Q or D → N mutations) or alter (K → D and D → K mutations) the salt-bridges in the matrix network. Proteins were recombinantly expressed in Escherichia coli membrane, reconstituted in model lipid membranes, and their structures and functions were analyzed by gel electrophoresis, circular dichroism spectroscopy, fluorescence assays, as well as molecular dynamics simulations. It is shown that the UCP2 matrix network contains five salt-bridges (rather than the previously reported three), and the matrix network can regulate the proton transport by holding the protein's transmembrane helices in close proximity, limiting the movement of the activator fatty acid(s). A biphasic two-state molecular model is proposed for proton transport in tetrameric (a dimer of stable dimers) UCP2, in which all the monomers are functional, and monomers in each dimer are in the same transport mode. Purine nucleotide (e.g., ATP) can occlude the internal pore of the monomeric units of UCP tetramers via interacting with positive residues at or in the proximity of the matrix network (K38, K141, K239, R88, R185, and R279) and prevent switching between cytoplasmic and matrix states, thus inhibiting the proton transport. This study provides new insights into the mechanism of proton transport and regulation in UCPs.

Functional Oligomeric Forms of Uncoupling Protein 2: Strong Evidence for Asymmetry in Protein and Lipid Bilayer Systems.

Stoichiometry of uncoupling proteins (UCPs) and their coexistence as functional monomeric and associated forms in lipid membranes remain intriguing open questions. In this study, tertiary and quaternary structures of UCP2 were analyzed experimentally and through molecular dynamics (MD) simulations. UCP2 was overexpressed in the inner membrane of Escherichia coli, then purified and reconstituted in lipid vesicles. Structure and proton transport function of UCP2 were characterized by circular dichroism (CD) spectroscopy and fluorescence methods. Findings suggest a tetrameric functional form for UCP2. MD simulations conclude that tetrameric UCP2 is a dimer of dimers, is more stable than its monomeric and dimeric forms, is asymmetrical and induces asymmetry in the membrane's lipid structure, and a biphasic on-off switch between the dimeric units is its possible mode of transport. MD simulations also show that the water density inside the UCP2 monomer is asymmetric, with the cytoplasmic side having a higher water density and a wider radius. In contrast, the structurally comparable adenosine 5'-diphosphate (ADP)/adenosine 5'-triphosphate (ATP) carrier (AAC1) did not form tetramers, implying that tetramerization cannot be generalized to all mitochondrial carriers.

Insights into the Polyhexamethylene Biguanide (PHMB) Mechanism of Action on Bacterial Membrane and DNA: A Molecular Dynamics Study.

Polyhexamethylene biguanide (PHMB) is a cationic polymer with antimicrobial and antiviral properties. It has been commonly accepted that the antimicrobial activity is due to the ability of PHMB to perforate the bacterial phospholipid membrane leading ultimately to its death. In this study, we show by the means of atomistic molecular dynamics (MD) simulations that, while the PHMB molecules attach to the surface of the phospholipid bilayer and partially penetrate it, they do not cause any pore formation at least within the microsecond simulation times. The polymers initially adsorb onto the membrane surface via the favorable electrostatic interactions between the phospholipid headgroups and the biguanide groups and then partially penetrate the membrane slightly disrupting its structure. This, however, does not lead to the formation of any pores. The microsecond-scale simulations reveal that it is unlikely for PHMB to spontaneously pass through the phospholipid membrane. Our findings suggest that PHMB translocation across the bilayer may take place through binding to the phospholipids. Once inside the cell, the polymer can effectively "bind" to DNA through extensive interactions with DNA phosphate backbone, which can potentially block the DNA replication process or activate DNA repair pathways.

Manipulation of Diatomic Molecules with Oriented External Electric Fields: Linear Correlations in Atomic Properties Lead to Nonlinear Molecular Responses.

Oriented external electric fields (OEEFs) have been shown to have great potential in being able to provide unprecedented control of chemical reactions, catalysis, and selectivity with applications ranging from H2 storage to molecular machines. We report a theoretical study of the atomic origins of molecular changes because of OEEFs since understanding the characteristics of OEEF-induced couplings between atomic and molecular properties is an important step toward comprehensive understanding of the effects of strong external fields on the molecular structure, stability, and reactivity. We focus on the atomic and molecular (bond) properties of a set of homo- (H2, N2, O2, F2, and Cl2) and heterodiatomic (HF, HCl, CO, and NO) molecules under intense external electric fields in the context of quantum theory of atoms in molecules (QTAIM). It is shown that the atomic properties (atomic charges, energies, and localization indices) correlate linearly with the field strengths, but molecular properties (bond length, electron density at the bond critical point, and electron delocalization index) exhibit nonlinear responses to the imposed fields. In particular, the changes in the electron density distribution alter the shapes and locations of the zero-flux surfaces, atomic volumes, atomic electron population, and localization/delocalization indices. The topography and topology of the molecular electrostatic potential undergo dramatic changes. External fields also perturb the covalent-polar-ionic characteristic of the studied chemical bonds, hallmarking the impact of electric fields on the stability and reactivity of chemical compounds. The findings are well-rationalized within the framework of the QTAIM and form a coherent conceptual understanding of these effects in prototypical diatomic molecules.

Membrane Disruption by Very Long Chain Fatty Acids during Necroptosis.

Necroptosis is a form of regulated cell death which results in loss of plasma membrane integrity, release of intracellular contents, and an associated inflammatory response. We previously found that saturated very long chain fatty acids (VLCFAs), which contain ≥20 carbons, accumulate during necroptosis. Here, we show that genetic knockdown of Fatty Acid (FA) Elongase 7 (ELOVL7) reduces accumulation of specific very long chain FAs during necroptosis, resulting in reduced necroptotic cell death and membrane permeabilization. Conversely, increasing the expression of ELOVL7 increases very long chain fatty acids and membrane permeabilization. In vitro, introduction of the VLCFA C24 FA disrupts bilayer integrity in liposomes to a greater extent than a conventional C16 FA. To investigate the microscopic origin of these observations, atomistic Molecular Dynamics (MD) simulations were performed. MD simulations suggest that fatty acids cause clear differences in bilayers based on length and that it is the interdigitation of C24 FA between the individual leaflets that results in disorder in the region and, consequently, membrane disruption. We synthesized clickable VLCFA analogs and observed that many proteins were acylated by VLCFAs during necroptosis. Taken together, these results confirm the active role of VLCFAs during necroptosis and point to multiple potential mechanisms of membrane disruption including direct permeabilization via bilayer disruption and permeabilization by targeting of proteins to cellular membranes by fatty acylation.

Manipulation of a DNA aptamer-protein binding site through arylation of internal guanine residues.

Chemically modified aptamers have the opportunity to increase aptamer target binding affinity and provide structure-activity relationships to enhance our understanding of molecular target recognition by the aptamer fold. In the current study, 8-aryl-2'-deoxyguanosine nucleobases have been inserted into the G-tetrad and central TGT loop of the thrombin binding aptamer (TBA) to determine their impact on antiparallel G-quadruplex (GQ) folding and thrombin binding affinity. The aryl groups attached to the dG nucleobase vary greatly in aryl ring size and impact on GQ stability (∼20 °C change in GQ thermal melting (Tm) values) and thrombin binding affinity (17-fold variation in dissociation constant (Kd)). At G8 of the central TGT loop that is distal from the aptamer recognition site, the probes producing the most stable GQ structure exhibited the strongest thrombin binding affinity. However, within the G-tetrad, changes to the electron density of the dG component within the modified nucleobase can diminish thrombin binding affinity. Detailed molecular dynamics (MD) simulations on the modified TBA (mTBA) and mTBA-protein complexes demonstrate how the internal 8-aryl-dG modification can manipulate the interactions between the DNA nucleobases and the amino acid residues of thrombin. These results highlight the potential of internal fluorescent nuclobase analogs (FBAs) to broaden design options for aptasensor development.

Structural Insight into the Discrimination between 8-Oxoguanine Glycosidic Conformers by DNA Repair Enzymes: A Molecular Dynamics Study of Human Oxoguanine Glycosylase 1 and Formamidopyrimidine-DNA Glycosylase.

hOgg1 and FPG are the primary DNA repair enzymes responsible for removing the major guanine (G) oxidative product, namely, 7,8-dihydro-8-oxoguanine (OG), in humans and bacteria, respectively. While natural G adopts the anti conformation and forms a Watson-Crick pair with cytosine (C), OG can also adopt the syn conformation and form a Hoogsteen pair with adenine (A). hOgg1 removes OG paired with C but is inactive toward the OG:A pair. In contrast, FPG removes OG from OG:C pairs and also exhibits appreciable (although diminished) activity toward OG:A pairs. As a first step toward understanding this difference in activity, we have employed molecular dynamics simulations to examine how the anti and syn conformers of OG are accommodated in the hOgg1 and FPG active sites. When anti-OG is bound, hOgg1 active site residues are properly aligned to initiate catalytic base departure, while geometrical parameters required for the catalytic reaction are not conserved for syn-OG. On the other hand, the FPG catalytic residues are suitably aligned for both OG conformers, with anti-OG being more favorably bound. Thus, our data suggests that the differential ability of hOgg1 and FPG to accommodate the anti- and syn-OG glycosidic conformations is an important factor that contributes to the relative experimental excision rates. Nevertheless, the positions of the nucleophiles with respect to the lesion in the active sites suggest that the reactant complex is poised to initiate catalysis through a similar mechanism for both repair enzymes and supports a recently proposed mechanism in which sugar-ring opening precedes nucleoside deglycosylation.

Quantum mechanical study of the ß- and δ-lyase reactions during the base excision repair process: application to FPG.

Bacterial FPG (or MutM) is a bifunctional DNA glycosylase that is primarily responsible for excising 8-oxoguanine (OG) from the genome by cleaving the glycosidic bond and the DNA backbone at the 3'- and 5'-phosphates of the damaged nucleoside. In the present work, quantum mechanical methods (SMD-M06-2X/6-311+G(2df,2p)//IEF-PCM-B3LYP/6-31G(d)) and a ring-opened Schiff base model that includes both the 3'- and 5'-phosphate groups are used to investigate the ß- and δ-elimination reactions facilitated by FPG. Both the ß- and δ-elimination reactions are shown to proceed through an E1cB mechanism that involves proton abstraction prior to the phosphate-ribose bond cleavage. Since transition states for the phosphate elimination reactions could not be characterized in the absence of leaving group protonation, our work confirms that the phosphate elimination reactions require protonation by a residue in the FPG active site, and can likely be further activated by additional active-site interactions. Furthermore, our model suggests that 5'-PO4 activation may proceed through a nearly isoenergetic direct (intramolecular) proton transfer involving the O4' proton of the deoxyribose of the damaged nucleoside. Regardless, our model predicts that both 3'- and 5'-phosphate protonation and elimination steps occur in a concerted reaction. Most importantly, our calculated barriers for the phosphate cleavage reactions reveal inherent differences between the ß- and δ-elimination steps. Indeed, our calculations provide a plausible explanation for why the δ-elimination rather than the ß-elimination is the rate-determining step in the BER facilitated by FPG, and why some bifunctional glycosylases (including the human counterpart, hOgg1) lack δ-lyase activity. Together, the new mechanistic features revealed by our work can be used in future large-scale modeling of the DNA-protein system to unveil the roles of key active sites residues in these relatively unexplored BER steps.

Lewis acid behavior of SF4 : synthesis, characterization, and computational study of adducts of SF4 with pyridine and pyridine derivatives.

Sulfur tetrafluoride was shown to act as a Lewis acid towards organic nitrogen bases, such as pyridine, 2,6-dimethylpyridine, 4-methylpyridine, and 4-dimethylaminopyridine. The SF4 ⋅NC5 H5 , SF4 ⋅2,6-NC5 H3 (CH3 )2 , SF4 ⋅4-NC5 H4 (CH3 ), and SF4 ⋅4-NC5 H4 N(CH3 )2 adducts can be isolated as solids that are stable below -45 °C. The Lewis acid-base adducts were characterized by low-temperature Raman spectroscopy and the vibrational bands were fully assigned with the aid of density functional theory (DFT) calculations. The electronic structures obtained from the DFT calculations were analyzed by the quantum theory of atoms in molecules (QTAIM). The crystal structures of SF4 ⋅NC5 H5 , SF4 ⋅4-NC5 H4 (CH3 ), and SF4 ⋅4-NC5 H4 N(CH3 )2 revealed weak SN dative bonds with nitrogen coordinating in the equatorial position of SF4 . Based on the QTAIM analysis, the non-bonded valence shell charge concentration on sulfur, which represents the lone pair, is only slightly distorted by the weak dative SN bond. No evidence for adducts between quinoline or isoquinoline with SF4 was found by low-temperature Raman spectroscopy.

Computational investigation of glycosylase and ß-lyase activity facilitated by proline: applications to FPG and comparisons to hOgg1.

Quantum mechanical methods are used to investigate the chemical steps during the bifunctional (glycosylase and ß-lyase) activity of bacterial FPG DNA glycosylase, which removes the major oxidation product (8-oxoguanine) from DNA as part of the base excision repair process. To facilitate investigation of all potential pathways, the smallest chemically relevant model is implemented, namely a modified OG nucleoside-3'-monophosphate and a truncated proline nucleophile. Potential energy surfaces are characterized with SMD-M06-2X/6-311+G(2df,2p)//PCM-B3LYP/6-31G(d) and compared to a previous study on the analogues human enzyme (hOgg1), which uses a lysine nucleophile (Kellie, J. L.; Wetmore, S. D. J. Phys. Chem. B 2012, 116, 10786-10797). Our large calculated barriers indicate that FPG must actively catalyze the three main phases of the overall reaction, namely, deglycosylation, (deoxyribose) ring-opening, and ß-elimination, and provide clues about how this is achieved through comparison to accurate crystal structures. The main conclusions about key mechanistic steps hold true regardless of the nucleophile, suggesting that most major differences in the relative activity of FPG and hOgg1 are primarily due to other active site residues. Nevertheless, support for possible monofunctional (deglycosylation only) activity is only evident when lysine is the nucleophile. This finding agrees with experimental observations of monofunctional activity of hOgg1 and further supports the broadly accepted bifunctional activity of FPG.

The chemical bond in external electric fields: energies, geometries, and vibrational Stark shifts of diatomic molecules.

It is shown that the response of molecular properties of diatomics such as the total energy, the bond length, and the vibrational Stark shift to an external homogenous electric field (EF) can be predicted from field-free observable properties such as the equilibrium bond length, the bond dissociation energy, the polarizability and dipole moment functions, and the vibrational frequency. Delley [J. Mol. Struct.: THEOCHEM 434, 229 (1998)] suggested to approximate the potential energy surface under an EF by a Morse function augmented with a EF term proportional to the internuclear separation. In this work, this term is replaced by the expression of the field-induced energy change which yields a field-perturbed Morse potential that tends to a constant asymptotic limit when the EF term itself become proportional to the sum of the polarizabilities of the separated atoms. The model is validated by comparison with direct calculations on nine diatomics, five homo-nuclear (H2, N2, O2, F2, and Cl2) and four hetero-nuclear (HF, HCl, CO, and NO), covering a range and combinations of dipole moments and polarizabilities. Calculations were conducted at the quadratic configuration interaction with single and double excitations (QCISD) and density functional theory (DFT)-B3LYP levels of theory using the 6-311++G(3df,2pd) basis set. All results agree closely at the two levels of theory except for the Stark effect of NO which is not correctly predicted by QCISD calculations as further calculations, including at the coupled cluster with single and double excitation (CCSD) level of theory, demonstrate.

Dipole moment surfaces of the CH4 + •X → CH3• + HX (X = F, Cl) reactions from atomic dipole moment surfaces, and the origins of the sharp extrema of the dipole moments near the transition states.

The partitioning of the dipole moment of an isolated molecule or that of a reacting system is reviewed and applied to a dynamic reacting system whereby the system's dipole moment surface is constructed in parallel to its potential energy surface. The dipole moment surface is then decomposed into two origin-independent surfaces: (1) an atomic polarization (AP) surface and a charge transfer (CT) surface. The dipole moment surface as well as its two composing AP and CT surfaces are all further broken down into atomic and/or group contributions with the aid of the quantum theory of atoms in molecules (QTAIM). This approach is applied to the title's laser-induced chemical reactions [CH4 + (•)X → CH3(•) + HX (X = F, Cl)] previously studied by Bandrauk et al. [ J. Chem. Phys. 2004 , 121 , 7764 - 7775 ], and which were found to exhibit marked peaks in the dipole moment and in the polarizability tensor component at (or near) the transition state. These peaks afford a means to control the kinetics of these reactions with the proper adjustment of an external laser field intensity and phase. The entrance channel potentials of these reactions have recently been probed by photodetachment spectroscopy by Bowman and collaborators [ J. Chem. Phys. 2011 , 134 , 191102_1 - 4 ]. The understanding of the origin of the peaks in the dipole moment can provide, eventually, an additional layer of control in the design of reactions tunable by external fields through the proper selection of the reactants to maximize the field-molecule interaction.