Performance of polarization-consistent vs. correlation-consistent basis sets for CCSD(T) prediction of water dimer interaction energy

Journal of Molecular Modeling - The chemical reactivity of the first- and second-generation Grubbs catalysts has always been a significant issue in olefin metathesis. In the present work, we study...     

β-turn tendency in N-methylated peptides with dehydrophenylalanine residue: DFT study

The tendency to adopt β‐turn conformation by model dipeptides with α,β‐dehydrophenylalanine (ΔPhe) residue in the gas phase and in solution is investigated by theoretical methods. We pay special attention to a dependence of conformational properties on the side‐chain configuration of dehydro residue and the influence of N‐methylation on β‐turn stability. An extensive computational study of the conformational preferences of Z and E isomers of dipeptides Ac‐Gly‐(E/Z)‐ΔPhe‐NHMe ( 1a / 1b ) and Ac‐Gly‐(E/Z)‐ΔPhe‐NMe₂ ( 2a / 2b ) by B3LYP/6‐311++G(d,p) and MP2/6‐311++G(d,p) methods is reported. It is shown that, in agreement with experimental data, Ac‐Gly‐(Z)‐ΔPhe‐NHMe has a great tendency to adopt β‐turn conformation. In the gas phase the type II β‐turn is preferred, whereas in the polar environment, the type I. On the other hand, dehydro residue in Ac‐Gly‐(E)‐ΔPhe‐NHMe has a preference to adopt extended conformations in all environments. N‐methylation of C‐terminal amide group, which prevents the formation of 1←4 intramolecular hydrogen bond, change dramatically the conformational properties of studied dehydropeptides. Especially, the tendency to adopt β‐turn conformations is much weaker for the N‐methylated Z isomer (Ac‐Gly‐(Z)‐ΔPhe‐NMe₂), both in vacuo and in the polar environment. On the contrary, N‐methylated E isomer (Ac‐Gly‐(E)‐ΔPhe‐NMe₂) can easier adopt β‐turn conformation, but the backbone torsion angles (?₁, ψ₁, ?₂, ψ₂) are off the limits for common β‐turn types. © 2012 Wiley Periodicals, Inc. Biopolymers 97:518–528, 2012.     

Thermodynamic characterization of binding Oxytricha nova single strand telomere DNA with the alpha protein N-terminal domain

The Oxytricha nova telemere binding protein alpha subunit binds single strand DNA and participates in a nucleoprotein complex that protects the very ends of chromosomes. To understand how the N-terminal, DNA binding domain of alpha interacts with DNA we measured the stoichiometry, enthalpy (DeltaH), entropy (DeltaS), and dissociation constant (K(D-DNA)) for binding telomere DNA fragments at different temperatures and salt concentrations using native gel electrophoresis and isothermal titration calorimetry (ITC). About 85% of the total free energy of binding corresponded with non-electrostatic interactions for all DNAs. Telomere DNA fragments d(T(2)G(4)), d(T(4)G(4)), d(G(3)T(4)G(4)), and d(G(4)T(4)G(4)) each formed monovalent protein complexes. In the case of d(T(4)G(4)T(4)G(4)), which has two tandemly repeated d(TTTTTGGGG) telomere motifs, two binding sites were observed. The high-affinity "A site" has a dissociation constant, K(D-DNA(A)) = 13(+/-4) nM, while the low-affinity "B site" is characterized by K(D-DNA(B)) = 5600(+/-600) nM at 25 degrees C. Nucleotide substitution variants verified that the A site corresponds principally with the 3'-terminal portion of d(T(4)G(4)T(4)G(4)). The relative contributions of entropy (DeltaS) and enthalpy (DeltaH) for binding reactions were DNA length-dependent as was heat capacity (DeltaCp). These trends with respect to DNA length likely reflect structural transitions in the DNA molecule that are coupled with DNA-protein association. Results presented here are important for understanding early intermediates and subsequent stages in the assembly of the full telomere nucleoprotein complex and how binding events can prepare the telomere DNA for extension by telomerase, a critical event in telomere biology.     

AIDS and associated malignancies

INTRODUCTION It has been over twenty years since the onset of the AIDS epidemic, and in spite of the tremendous progress made towards the understanding of the disease, the virus that causes the disease and the development of highly ef- fective anti-retrov…     

Total chemical synthesis and oxidative folding of delta-conotoxin PVIA containing an N-terminal propeptide

Small disulfide-rich peptides are translated as larger precursors typically containing an N-terminal prepro sequence. In this study, we investigated the role of a propeptide in the oxidative folding of an extremely hydrophobic delta-conotoxin, PVIA. delta-Conotoxin PVIA (delta-PVIA) is a 29-amino acid neurotoxin stabilized by three disulfide bridges. Previous folding studies on delta-conotoxins revealed that their poor folding properties resulted from their hydrophobicity. However, low folding yields of delta-PVIA could be improved by the presence of a nonionic detergent, which acted as a chemical chaperone. delta-PVIA provided an attractive model to investigate whether the hydrophilic propeptide region could function as an intramolecular chaperone. A 58-amino acid precursor for delta-PVIA (pro-PVIA), containing the N-terminal propeptide covalently attached to the mature conotoxin, was synthesized using native chemical ligation. Oxidative folding of pro-PVIA resulted in a very low accumulation of the correctly folded form, comparable to that for the mature conotoxin delta-PVIA. Our results are in accord with the relevant data previously observed for alpha- and omega-conotoxins, indicating that conotoxin prepro sequences are so-called class II propeptides, which are not directly involved in the oxidative folding. We hypothesize that these propeptide regions may be important for interactions with protein folding catalysts and sorting receptors during the secretory process.     

Propeptide does not act as an intramolecular chaperone but facilitates protein disulfide isomerase-assisted folding of a conotoxin precursor

Conotoxins comprise a large and diverse group of peptide neurotoxins derived from Conus snail venoms; most contain multiple disulfide bonds. The conotoxin precursors consist of three distinct domains: the N-terminal signal sequence, an intervening propeptide region, and the C-terminal mature conotoxin. Formation of the native disulfide bonds during the oxidative folding of conotoxins is a prerequisite for their proper biological function, but in numerous in vitro folding experiments with mature conotoxins, a lack of specificity in formation of the native Cys-Cys connectivities is observed. The mechanisms that ensure that the native disulfide bonds are formed in venom ducts during biosynthesis remain unknown. To evaluate whether the propeptide could potentially function as an intramolecular chaperone, we studied the oxidative folding of a conotoxin precursor, pro-GI, belonging to the alpha-conotoxin family. Our results indicate that the propeptide sequence did not directly contribute to folding kinetics and thermodynamics. However, we found that the propeptide region of pro-GI played an important role when oxidative folding was catalyzed by protein disulfide isomerase (PDI). The PDI-assisted reaction was more efficient during the early folding in the context of the propeptide sequence (pro-GI), as compared to that of the mature conotoxin (alpha-GI). Taken together, our results suggest for the first time that the propeptide region may play a role in the PDI-catalyzed oxidative folding of conotoxin precursors.     

The effect of Cu2+ on uptake and assimilation of ammonium by cucumber seedlings

The ammonium uptake by cucumber seedlings was estimated from ammonium ions depletion in an uptake solution. The uptake of NH ₄ ⁺ was decreased by about 60 % after one hour and by about 90 % after two hours of 100 μM Cu²⁺ treatment. On the contrary the accumulation of ammonium in roots of Cu²⁺-treated seedlings at the same time was higher than in the control. Cu²⁺ in the concentration inhibiting NH ₄ ⁺ absorption during one hour inhibited also glutamine synthetase (GS) (EC 6.3.1.2) and NADH-glutamate dehydrogenase (NADH-GDH) (EC 1.4.1.2) activities both localized in the roots of seedlings. After one hour and at least up to the 4th hour Cu²⁺ accumulated mainly in roots (95 %). It was probably the reason of the GS activity in cotyledons of seedling treated with Cu²⁺ that it was at the same level as in the control. NADH-GDH activity in cotylcdons after one hour of the Cu²⁺ treatment was lower than in the control but the influence of Cu²⁺ action on the activity of this enzyme in roots was by far stronger. 100 μM Cu²⁺ did not affect the activities of both enzymes in in vitro experiments. Copper added into the incubation medium in 1000 μM concentration decreased GS activity, but still did not change NADH-GDH activity. These results suggested the indirect Cu²⁺ action on the investigated enzymes in in vivo experiments. However, no substantial effect on enzyme activities extracted from control plants was observed after the addition of the extract from Cu²⁺-treated plants into the incubation medium. The data suggest that the influence of Cu²⁺ on uptake and assimilation of ammonium may be connected not only with changes of plasma membrane properties in the root cells of Cu²⁺ treated seedlings but also with Cu²⁺ action on two major enzymes involved in NH ₄ ⁺ assimilation: glutamate synthetase and NADH-glutamate dehydrogenase.     

Characterization of the nitrate transporter in root plasma membranes of Cucumis sativus L

Nitrate uptake in right-side out plasma membrane vesicles isolated from cucumber roots was characterized. Nitrate uptake into vesicles was driven by an artificially imposed pH gradient. The uptake was strongly inhibited by phenylglyoxal, an arginyl residue modificator. Only a slight repression of NO ₃ ⁻ transport in vesicles was observed in the presence of NEM, a thiol group reagent. pCMBS, an other thiol reagent and DEPC, an effector of histidine residue, had no effect on the nitrate transport in plasma membranes. ATP-driven proton transport in vesicles was not significantly affected in the presence of both, phenylglyoxal and DEPC, whereas pCMBS and NEM abolished it almost completely. The possible role of the particular amino acids residues in the active nitrate transport is discussed. NO ₃ ⁻ uptake into vesicles isolated from both, nitrate-induced and nitrate-depleted plant material was higher than that observed in the vesicles obtained from uninduced plants. Thus, isolated vesicles reflect the well-known in vivo response of intact plants on the exogenous nitrogen regime.