Theoretical Studies of Furan and Thiophene Nanothreads: Structures, Cycloaddition Barriers and Activation Volumes

<p>In this theoretical study we examine several aspects of the formation, structure, and stability of the most ordered nanothreads yet made, those derived from furan and thiophene. First, we look at the enthalpic consequences and activation barriers of the first two steps of oligomerization by a Diels-Alder mechanism. The ca. 20 GPa difference in the synthetic pressures (furan lower) is explainable in terms of greater loss of aromaticity by the thiophene. Subsequent steps have understandably lower barriers. We show explicitly how pressure affects the reaction profiles, operating through the volume decrease in the transition state and onward to the product molecule. The interesting option of polymerization proceeding in one or two directions opens up the possibility of polymers with two opposing and cumulative dipole moments. The computed activation volumes are consistently more negative for likely initial furan (compared with thiophene) polymerization steps, in accord with the lower onset pressure of furan polymerization. In the second part of our study we examine the energetics of the likely polymers. Three ordered polymer structures compete in enthalpy -- a <i>syn</i> one, with all O/S on the same side, an <i>anti </i>one, S/O alternating, and a<i> syn-anti</i> isomer, with segments of four monomers repeating. The <i>syn</i> polymer, if not allowed to distort, is at high enthalpy relative to the other two. The origin of the destabilization is apparent, being S/O lone-pair repulsion, understandably greater for S than O at the 2.8/2.6Å separation. Set free, the <i>syn</i> isomers curve or arc, in two- or three-dimensional (helical) ways, whose energetics are traced in detail. The <i>syn</i> polymer can also stabilize itself by the thread twisting into zig-zag or helical enthalpic minima. Release of strain in a linear thread as the pressure is relaxed to 1 atm, with consequent thread curving, is a likely mechanism for the observed loss of crystalline order in the polymer as it is returned to ambient pressure.<br></p>

Theoretical Studies of Furan and Thiophene Nanothreads: Structures, Cycloaddition Barriers and Activation Volumes

<p>In this theoretical study we examine several aspects of the formation, structure, and stability of the most ordered nanothreads yet made, those derived from furan and thiophene. First, we look at the enthalpic consequences and activation barriers of the first two steps of oligomerization by a Diels-Alder mechanism. The ca. 20 GPa difference in the synthetic pressures (furan lower) is explainable in terms of greater loss of aromaticity by the thiophene. Subsequent steps have understandably lower barriers. We show explicitly how pressure affects the reaction profiles, operating through the volume decrease in the transition state and onward to the product molecule. The interesting option of polymerization proceeding in one or two directions opens up the possibility of polymers with two opposing and cumulative dipole moments. The computed activation volumes are consistently more negative for likely initial furan (compared with thiophene) polymerization steps, in accord with the lower onset pressure of furan polymerization. In the second part of our study we examine the energetics of the likely polymers. Three ordered polymer structures compete in enthalpy -- a <i>syn</i> one, with all O/S on the same side, an <i>anti </i>one, S/O alternating, and a<i> syn-anti</i> isomer, with segments of four monomers repeating. The <i>syn</i> polymer, if not allowed to distort, is at high enthalpy relative to the other two. The origin of the destabilization is apparent, being S/O lone-pair repulsion, understandably greater for S than O at the 2.8/2.6Å separation. Set free, the <i>syn</i> isomers curve or arc, in two- or three-dimensional (helical) ways, whose energetics are traced in detail. The <i>syn</i> polymer can also stabilize itself by the thread twisting into zig-zag or helical enthalpic minima. Release of strain in a linear thread as the pressure is relaxed to 1 atm, with consequent thread curving, is a likely mechanism for the observed loss of crystalline order in the polymer as it is returned to ambient pressure.<br></p>

Theoretical Studies of Furan and Thiophene Nanothreads: Structures, Cycloaddition Barriers and Activation Volumes

<p>In this theoretical study we examine several aspects of the formation, structure, and stability of the most ordered nanothreads yet made, those derived from furan and thiophene. First, we look at the enthalpic consequences and activation barriers of the first two steps of oligomerization by a Diels-Alder mechanism. The ca. 20 kcal/mol difference in the synthetic pressures (furan lower) is explainable in terms of greater loss of aromaticity by the thiophene. Subsequent steps have understandably lower barriers. We show explicitly how pressure affects the reaction profiles, operating through the volume decrease in the transition state and onward to the product molecule. The interesting option of polymerization proceeding in one or two directions opens up the possibility of polymers with two opposing and cumulative dipole moments. The computed activation volumes are consistently more negative for likely initial furan (compared with thiophene) polymerization steps, in accord with the lower onset pressure of furan polymerization. In the second part of our study we examine the energetics of the likely polymers. Three ordered polymer structures compete in enthalpy -- a <i>syn</i> one, with all O/S on the same side, an <i>anti </i>one, S/O alternating, and a<i> syn-anti</i> isomer, with segments of four monomers repeating. The <i>syn</i> polymer, if not allowed to distort, is at high enthalpy relative to the other two. The origin of the destabilization is apparent, being S/O lone-pair repulsion, understandably greater for S than O at the 2.8/2.6Å separation. Set free, the <i>syn</i> isomers curve or arc, in two- or three-dimensional (helical) ways, whose energetics are traced in detail. The <i>syn</i> polymer can also stabilize itself by the thread twisting into zig-zag or helical enthalpic minima. Release of strain in a linear thread as the pressure is relaxed to 1 atm, with consequent thread curving, is a likely mechanism for the observed loss of crystalline order in the polymer as it is returned to ambient pressure.<br></p>

Finding metabolic pathways in large networks through atom-conserving substrate-product pairs

Finding biosynthetic pathways is essential for metabolic engineering of organisms to produce chemicals, biodegradation prediction of pollutants and drugs, and for the elucidation of bioproduction pathways of secondary metabolites. A key step in biosynthetic pathway design is the extraction of novel metabolic pathways from big networks that integrate known biological, as well as novel, predicted biotransformations. However, especially with the integration of big data, the efficient analysis and navigation of metabolic networks remains a challenge. Here, we propose the construction of searchable graph representations of metabolic networks. Éach reaction is decomposed into pairs of reactants and products, and each pair is assigned a weight, which is calculated from the number of conserved atoms between the reactant and the product molecule. We test our method on a biochemical network that spans 6,546 known enzymatic reactions to show how our approach elegantly extracts biologically relevant metabolic pathways from biochemical networks, and how the proposed network structure enables the application of efficient graph search algorithms that improve navigation and pathway identification in big metabolic networks. The weighted reactant-product pairs of an example network and the corresponding graph search algorithm are available online. The proposed method extracts metabolic pathways fast and reliably from big biochemical networks, which is inherently important for all applications involving the engineering of metabolic networks.

<i>Geminal</i> parahydrogen-induced polarization: accumulating long-lived singlet order on methylene proton pairs

In the majority of hydrogenative parahydrogen-induced polarization (PHIP) experiments, the hydrogen molecule undergoes pairwise cis addition to an unsaturated precursor to occupy vicinal positions on the product molecule. However, some ruthenium-based hydrogenation catalysts induce geminal hydrogenation, leading to a reaction product in which the two hydrogen atoms are transferred to the same carbon centre, forming a methylene (CH2) group. The singlet order of parahydrogen is substantially retained over the geminal hydrogenation reaction, giving rise to a singlet-hyperpolarized CH2 group. Although the T1 relaxation times of the methylene protons are often short, the singlet order has a long lifetime, provided that singlet–triplet mixing is suppressed, either by chemical equivalence of the protons or by applying a resonant radiofrequency field. The long lifetime of the singlet order enables the accumulation of hyperpolarization during the slow hydrogenation reaction. We introduce a kinetic model for the behaviour of the observed hyperpolarized signals, including both the chemical kinetics and the spin dynamics of the reacting molecules. Our work demonstrates the feasibility of producing singlet-hyperpolarized methylene moieties by parahydrogen-induced polarization. This potentially extends the range of molecular agents which may be generated in a hyperpolarized state by chemical reactions of parahydrogen.

<i>Geminal</i> Parahydrogen-Induced Polarization: Accumulating Long-Lived Singlet Order on Methylene Proton Pairs

In the majority of hydrogenative PHIP (Parahydrogen Induced Polarization) experiments, the hydrogen molecule undergoes pairwise cis-addition to an unsaturated precursor to occupy vicinal positions on the product molecule. However, some ruthenium-based hydrogenation catalysts induce geminal hydrogenation, leading to a reaction product in which the twohydrogen atoms are transferred to the same carbon center, forming a methylene (CH2) group. The singlet order of parahydrogen is substantially retained over the geminal hydrogenation reaction, giving rise to a singlet-hyperpolarized CH2 group. Although the T1 relaxation times of the methylene protons are often short, the singlet order has a long lifetime, providing that singlet-triplet mixing is suppressed, either by chemical equivalence of the protons or by applying a resonant radiofrequency field. The long lifetime of the singlet order enables the accumulation of hyperpolarization during the slow hydrogenation reaction. We introduce a kinetic model for the behaviour of the observed hyperpolarized signals, including both the chemical kinetics and the spin dynamics of the reacting molecules. Our work demonstrates the feasibility of producing singlet-hyperpolarized methylene moieties by parahydrogen-induced polarization. This potentially extends the range of molecular agents which maybe generated in a hyperpolarized state by chemical reactions of parahydrogen.

Ketoreductase TpdE fromRhodococcus jostiiTMP1: characterization and application in the synthesis of chiral alcohols

Background.Production of highly pure enantiomers of vicinal diols is desirable, but difficult to achieve. Enantiomerically pure diols and acyloins are valuable bulk chemicals, promising synthones and potential building blocks for chiral polymers. Enzymatic reduction of ketones is a useful technique for the synthesis of the desired enantiomeric alcohols. Here, we report on the characterization of a ketoreductase TpdE fromRhodococcus jostiiTMP1 that is a prospective tool for the synthesis of such compounds.Results.In this study, NADPH-dependent short-chain dehydrogenase/reductase TpdE fromRhodococcus jostiiTMP1 was characterized. The enzyme exhibited broad substrate specificity towards aliphatic 2,3-diketones, butan-3-one-2-yl alkanoates, as well as acetoin and its acylated derivatives. TpdE stereospecifically reducedα-diketones to the corresponding diols. The GC-MS analysis of the reduction products of 2,3- and 3,4-diketones indicated that TpdE is capable of reducing both keto groups in its substrate leading to the formation of two new chiral atoms in the product molecule. Bioconversions of diketones to corresponding diols occurred using either purified enzyme or a whole-cellEscherichia coliBL21 (DE3) biocatalyst harbouring recombinant TpdE. The optimum temperature and pH were determined to be 30–35 °C and 7.5, respectively.Conclusions.The broad substrate specificity and stereoselectivity of TpdE fromRhodococcus jostiiTMP1 make it a promising biocatalyst for the production of enantiomerically pure diols that are difficult to obtain by chemical routes.

Site-Directed Mutagenesis of N5-Carboxyaminoimidazole Ribonucleotide Mutase (Class I PurE) in Bacillus anthracis

Anthrax, the infection caused by the Gram-positive pathogen Bacillus anthracis (B. anthracis), is fatal if untreated, and some strains of B. anthracis have been found to be resistant to currently available antibiotics. The development of broad spectrum antibiotics is needed to treat the resistive strains. In antibiotic development, we have targeted B. anthracis Class I PurE enzyme (Ba PurE) as a unique and necessary enzyme in the de novo purine biosynthesis pathway, since the inactivation of this gene prevents B. anthraci growth in human serum, resulting in decreased bacterial proliferation. To identify inhibitors to $Ba$PurE, structural information on the substrate binding to its active site is needed. However, it is difficult to obtain crystals of Ba PurE with the substrate molecule in its binding site since upon binding to PurE, the substrate molecule is converted to the product molecule. An alternative approach is to create mutants of PurE that exhibit no enzymatic activity and do not convert the substrate to product, but still allow the substrate to bind to the active site. Then, the structure of mutant PurE with bound substrate can be obtained. We have identified a histidine residue at position 70 as the target of mutation to give an inactive enzyme. After successfully preparing the recombinant protein H70N, we have found that it exhibited no enzyme activity. This mutant will be useful in future experimentation to identify inhibitors of Ba PurE.

Theoretical study of the stereo-dynamics of the H + HeH+(v = 0, j = 0) → H2+ + He reaction

Theoretical study of the stereo-dynamics of the reaction, H + HeH+ (v = 0, j = 0) → H 2+ + He, have been performed with quasi-classical trajectory (QCT) method at different collision energies on a new ab initio potential energy surface. The distributions of P(θr), P(ϕr) and four generalized polarization-dependent differential cross-sections have been calculated. The results indicate that both the orientation and alignment of the rotational angular momentum are impacted by collision energies. With the collision energy increases, the rotation of the product molecule has a preference of changing from the “in-plane” reaction mechanism to the “out-of-plane” mechanism. Although the reaction is mainly dominated by the direct reaction mechanism, the indirect mechanism plays a role while the collision energy is very low.

Free and ATP-bound structures of Ap4A hydrolase fromAquifex aeolicusV5

Asymmetric diadenosine tetraphosphate (Ap4A) hydrolases degrade the metabolite Ap4A back into ATP and AMP. The three-dimensional crystal structure of Ap4A hydrolase (16 kDa) fromAquifex aeolicushas been determined in free and ATP-bound forms at 1.8 and 1.95 Å resolution, respectively. The overall three-dimensional crystal structure of the enzyme shows an αβα-sandwich architecture with a characteristic loop adjacent to the catalytic site of the protein molecule. The ATP molecule is bound in the primary active site and the adenine moiety of the nucleotide binds in a ring-stacking arrangement equivalent to that observed in the X-ray structure of Ap4A hydrolase fromCaenorhabditis elegans. Binding of ATP in the active site induces local conformational changes which may have important implications in the mechanism of substrate recognition in this class of enzymes. Furthermore, two invariant water molecules have been identified and their possible structural and/or functional roles are discussed. In addition, modelling of the substrate molecule at the primary active site of the enzyme suggests a possible path for entry and/or exit of the substrate and/or product molecule.