Microscopic simulation of quantum dynamics and nuclear tunneling in bacterial reaction centers

The dispersed polaron version of the semiclassical trajectory approach is used to evaluate the quantum mechanical nuclear tunneling effects in the charge recombination reaction, P⁺Q^–PQ, in photosynthetic bacterial reaction centers, The cclculations are based on the crystallographic structure of reaction centers from Rhodopseudomonas viridis. They succeed in capturing the temperature dependence of the rate constant without using adjustable parameters. This provides the first example of a microscopic simulation of quantum mechanical nuclear tunneling in a biological system.Abbreviations P bacteriochlorophyll dimer - Q ubiquinone in Rhodobacter sphaeroides, menaquinone in Rhodopseudomonas viridis - Rps. Rhodopseudomonas - Rb. Rhodobacter - QCFF/PI quantum mechanical extension of the consistent force field to -electron     

Sensitivity to plating of Escherichia coli cells expressing the cryA gene from Bacillus thuringiensis var. israelensis

Summary The gene (cytA) coding for the 27 kDa polypeptide of the Bacillus thuringiensis var. israelensis mosquito larvicidal -endotoxin, was cloned into a plasmid containing the T7 bacteriophage promoter. The plasmid was used to transform an Escherichia coli strain containing the T7 RNA polymerase gene 1, under the control of lacP. Loss of colony-forming ability without substantial lysis, associated with immediate inhibition of DNA synthesis, was observed after induction of transformed cells. The cytA gene product may kill E. colicells by disrupting their chromosome replicating apparatus.     

New withanolides from a cross of a South African chemotype by chemotype II (Israel) in Withania somnifera

Three new withanolides have been isolated from hybrids obtained by crossing a chemotype of Withania somnifera received from South Africa and chemotype II originating in Israel. The compounds have been characterized as 4β,20α-dihydroxy-1-oxo-5β,6β-epoxy-20R,22R-witha-24-enolide, 20α-hydroxy-1,4-dioxo-5β,6β-epoxy-20R,22R-witha-2,24-dienolide, and 20α-hydroxy-1,4-dioxo-5β,6β-epoxy-20R,22R-witha-2-enolide. The major steroid of the plant is withanolide D, while the other known withanolides present are 4β,20α-dihydroxy-1-oxo-5β,6β-epoxy-20R,22R,24S,25R-witha-2-enolide and withaferin A. The structures assigned to the new compounds are based on spectral evidence, analysis of their fragmentation under electron impact, and on chemical correlation with known compounds. The formation of these withanolides in this new hybrid is discussed briefly.     

Folding and stability of helical proteins: Carp myogen

In this work we use our very simple general representation of protein structures to study the mainly helical protein carp myogen. The representation, which treats the amino acid side-chains as simple spheres, is further simplified by rigidly fixing residues in α-helices. With this model we are able to reproduce the geometry and energetic stability of the native myogen conformation. Studies of the formation of α-helical sub-assemblies showed that the simulated folding of two and four-helix systems worked well, reaching compact native-like conformations with a good rate of success. Greater problems were encountered with the whole molecule (six helices), possibly due to the omission of entropic effects or to simulating the folding too rapidly. Finally, studies of the conformation of a pair of helices when isolated and when part of the whole molecule native conformation showed that long-range interactions have an unexpectedly strong influence on the conformation of the pair of helices.     

Associations between Vitamin D Status and Type 2 Diabetes Measures among Inuit in Greenland May Be Affected by Other Factors

ObjectiveEpidemiological studies have provided evidence of an association between vitamin D insufficiency and type 2 diabetes. Vitamin D levels have decreased among Inuit in Greenland, and type 2 diabetes is increasing. We hypothesized that the decline in vitamin D could have contributed to the increase in type 2 diabetes, and therefore investigated associations between serum 25(OH)D3 as a measure of vitamin D status and glucose homeostasis and glucose intolerance in an adult Inuit population.Methods2877 Inuit (≥18 years) randomly selected for participation in the Inuit Health in Transition study were included. Fasting- and 2hour plasma glucose and insulin, C-peptide and HbA_1c were measured, and associations with serum 25(OH)D3 were analysed using linear and logistic regression. A subsample of 330 individuals who also donated a blood sample in 1987, were furthermore included.ResultsAfter adjustment, increasing serum 25(OH)D3 (per 10 nmol/L) was associated with higher fasting plasma glucose (0.02 mmol/L, p = 0.004), 2hour plasma glucose (0.05 nmol/L, p = 0.002) and HbA_1c (0.39%, p<0.001), and with lower beta-cell function (-1.00 mmol/L, p<0.001). Serum 25(OH)D3 was positively associated with impaired fasting glycaemia (OR: 1.08, p = 0.001), but not with IGT or type 2 diabetes.ConclusionsOur results did not support an association between low vitamin D levels and risk of type 2 diabetes. Instead, we found weak positive associations between vitamin D levels and fasting- and 2hour plasma glucose levels, HbA_1c and impaired fasting glycaemia, and a negative association with beta-cell function, underlining the need for determination of the causal relationship.     

The control of the discrimination between dNTP and rNTP in DNA and RNA polymerase

Understanding the origin of discrimination between rNTP and dNTP by DNA/RNA polymerases is important both for gaining fundamental knowledge on the corresponding systems and for advancing the design of specific drugs. This work explores the nature of this discrimination by systematic calculations of the transition state (TS) binding energy in RB69 DNA polymerase (gp43) and T7 RNA polymerase. The calculations reproduce the observed trend, in particular when they included the water contribution obtained by the water flooding approach. Our detailed study confirms the idea that the discrimination is due to the steric interaction between the 2′OH and Tyr416 in DNA polymerase, while the electrostatic interaction is the source of the discrimination in RNA polymerase. Proteins 2016; 84:1616–1624. © 2016 Wiley Periodicals, Inc.     

Exploring the mechanism of DNA polymerases by analyzing the effect of mutations of active site acidic groups in Polymerase β

Elucidating the catalytic mechanism of DNA polymerase is crucial for a progress in the understanding of the control of replication fidelity. This work tries to advance the mechanistic understanding by analyzing the observed effect of mutations of the acidic groups in the active site of Polymerase β as well as the pH effect on the rate constant. The analysis involves both empirical valence bond (EVB) free energy calculations and considerations of the observed pH dependence of the reaction. The combined analysis indicates that the proton transfer (PT) from the nucleophilic O3′ has two possible pathways, one to D256 and the second to the bulk. We concluded based on calculations and the experimental pH profile that the most likely path for the wild‐type (WT) and the D256E and D256A mutants is a PT to the bulk, although the WT may also use a PT to Asp 256. Our analysis highlights the need for very extensive sampling in the calculations of the activation barrier and also clearly shows that ab initio QM/MM calculations that do not involve extensive sampling are unlikely to give a clear quantitative picture of the reaction mechanism. Proteins 2016; 84:1644–1657. © 2016 Wiley Periodicals, Inc.     

The catalytic effect of dihydrofolate reductase and its mutants is determined by reorganization energies

The effect of distant mutations on the catalytic reaction of dihydrofolate reductase (DHFR) is reexamined by empirical valence bond simulations. The simulations reproduce for the first time the changes in the observed rate constants (without the use of adjustable parameters for this purpose) and show that the changes in activation barriers are strongly correlated with the corresponding changes in the reorganization energy. The preorganization of the polar groups of enzymes is the key catalytic factor, and anticatalytic mutations destroy this preorganization. Some anticatalytic mutations in DHFR also increase the distance between the donor and acceptor, but this effect is not directly related to catalysis since the native enzyme and the uncatalyzed reaction in water have similar average donor-acceptor distances. Insight into the effect of a mutation is provided by constructing the relevant free energy surfaces in terms of the generalized solute-solvent coordinates. It is shown how the mutations change the reaction coordinate and the activation barrier, and it is clarified that the corresponding changes do not reflect dynamical effects. It is also pointed out that all reactions in a condensed phase involve correlated motions (both in enzymes and in solution) and that the change of such motions upon mutations is a result of the change in the shape of the multidimensional reaction path on the solute-solvent surface, rather than the reason for the change in rate constant. Thus, as far as catalysis is concerned, the change in the activation barrier is due to the change in the electrostatic preorganization energy.