Deamination of vasopressin (AVP) enhances its antidiuretic activity. Moreover, introduction of D-Arg8 instead of its L enantiomer in deamino-vasopressin (dAVP) results in an extremely potent and selective antidiuretic agonist - desmopressin (dDAVP). In this study we describe the synthesis, pharmacological properties and structures of these two potent antidiuretic agonists, and their inverso analogs. The structures of the peptides are studied in micellar and liposomic models of cell membrane using CD spectroscopy. Additionally, three-dimensional structures in mixed anionic-zwitterionic micelles are obtained using NMR spectroscopy supported by molecular dynamics simulations. Our conformational studies have shown that desmopressin in a membrane mimicking environment adopts one of the characteristic for vasopressin-like peptides β-turn - in position 3,4. Furthermore, dDAVP shows the tendency to create a β-turn in the Cys6-Gly9 C-tail, considered to be important for the antidiuretic activity, and also some tendency to adopt a 5,6 β-turn. In desmopressin, in contrast to the native vasopressin, deamino-vasopressin and [D-Arg8]-vasopressin (DAVP), the Arg8 side chain, crucial for the pressor and antidiuretic activities, is very well exposed for interaction with the receptor, whereas Gly9, crucial for the pressor and uterotonic activities, is situated together with the C-terminal amide group very close to the tocin ring. The arrangements of the Gln4 and Asn5 side chains, being crucial for OT activity, also differ in desmopressin as compared to those of AVP, dAVP and DAVP. These differences in arrangement of the important for activities side chains are likely to explain extremely potent and selective antidiuretic activities of desmopressin. © 2016 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 245-259, 2016.
- MeSH
- 1,2-dipalmitoylfosfatidylcholin chemie MeSH
- antidiuretika chemická syntéza farmakologie MeSH
- cyklizace MeSH
- desmopresin chemická syntéza farmakologie MeSH
- fluoreny chemie MeSH
- fosfatidylglyceroly chemie MeSH
- liposomy chemie MeSH
- micely MeSH
- nukleární magnetická rezonance biomolekulární MeSH
- potkani Wistar MeSH
- sekundární struktura proteinů MeSH
- simulace molekulární dynamiky MeSH
- techniky syntézy na pevné fázi metody MeSH
- uterotonika chemická syntéza farmakologie MeSH
- uterus účinky léků fyziologie MeSH
- vodíková vazba MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Repetitive extragenic palindrome (REP)-associated tyrosine transposase enzymes (RAYTs) bind REP DNA domains and catalyze their cleavage. Genomic sequence analyses identify potential noncoding REP sequences associated with RAYT-encoding genes. To probe the conformational space of potential RAYT DNA binding domains, we report here spectroscopic and calorimetric measurements that detect and partially characterize the solution conformational heterogeneity of REP oligonucleotides from six bacterial species. Our data reveal most of these REP oligonucleotides adopt multiple conformations, suggesting that RAYTs confront a landscape of potential DNA substrates in dynamic equilibrium that could be selected, enriched, and/or induced via differential binding. Thus, the transposase-bound DNA motif may not be the predominant conformation of the isolated REP domain. Intriguingly, for several REPs, the circular dichroism spectra suggest guanine tetraplexes as potential alternative or additional RAYT recognition elements, an observation consistent with these REP domains being highly nonrandom, with tetraplex-favoring 5'-G and 3'-C-rich segments. In fact, the conformational heterogeneity of REP domains detected and reported here, including the formation of noncanonical DNA secondary structures, may reflect a general feature required for recognition by RAYT transposases. Based on our biophysical data, we propose guanine tetraplexes as an additional DNA recognition element for binding by RAYT transposase enzymes.
In numerous Gram-positive bacteria, the glmS ribozyme or catalytic riboswitch regulates the expression of glucosamine-6-phosphate (GlcN6P) synthase via site-specific cleavage of its sugar-phosphate backbone in response to GlcN6P ligand binding. Biochemical data have suggested a crucial catalytic role for an active site guanine (G40 in Thermoanaerobacter tengcongensis, G33 in Bacillus anthracis). We used hybrid quantum chemical/molecular mechanical (QM/MM) calculations to probe the mechanism where G40 is deprotonated and acts as a general base. The calculations suggest that the deprotonated guanine G40(-) is sufficiently reactive to overcome the thermodynamic penalty arising from its rare protonation state, and thus is able to activate the A-1(2'-OH) group toward nucleophilic attack on the adjacent backbone. Furthermore, deprotonation of A-1(2'-OH) and nucleophilic attack are predicted to occur as separate steps, where activation of A-1(2'-OH) precedes nucleophilic attack. Conversely, the transition state associated with the rate-determining step corresponds to concurrent nucleophilic attack and protonation of the G1(O5') leaving group by the ammonium moiety of the GlcN6P cofactor. Overall, our calculations help to explain the crucial roles of G40 (as a general base) and GlcN6P (as a general acid) during glmS ribozyme self-cleavage. In addition, we show that the QM/MM description of the glmS ribozyme self-cleavage reaction is significantly more sensitive to the size of the QM region and the quality of the QM-MM coupling than that of other small ribozymes.
For mimicking macromolecular crowding of DNA quadruplexes, various crowding agents have been used, typically PEG, with quadruplexes of micromolar strand concentrations. Thermal and thermodynamic stabilities of these quadruplexes increased with the concentration of the agents, the rise depended on the crowder used. A different phenomenon was observed, and is presented in this article, when the crowder was the quadruplex itself. With DNA strand concentrations ranging from 3 µM to 9 mM, the thermostability did not change up to ∼2 mM, above which it increased, indicating that the unfolding quadruplex units were not monomolecular above ∼2 mM. The results are explained by self-association of the G-quadruplexes above this concentration. The ΔG(°) 37 values, evaluated only below 2 mM, did not become more negative, as with the non-DNA crowders, instead, slightly increased. Folding topology changed from antiparallel to hybrid above 2 mM, and then to parallel quadruplexes at high, 6-9 mM strand concentrations. In this range, the concentration of the DNA phosphate anions approached the concentration of the K(+) counterions used. Volume exclusion is assumed to promote the topological changes of quadruplexes toward the parallel, and the decreased screening of anions could affect their stability.
- MeSH
- cirkulární dichroismus MeSH
- denzitometrie MeSH
- DNA chemie MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- entropie MeSH
- G-kvadruplexy * MeSH
- lidé MeSH
- mikroskopie atomárních sil MeSH
- spektrofotometrie ultrafialová MeSH
- telomery chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Base stacking is a major interaction shaping up and stabilizing nucleic acids. During the last decades, base stacking has been extensively studied by experimental and theoretical methods. Advanced quantum-chemical calculations clarified that base stacking is a common interaction, which in the first approximation can be described as combination of the three most basic contributions to molecular interactions, namely, electrostatic interaction, London dispersion attraction and short-range repulsion. There is not any specific π-π energy term associated with the delocalized π electrons of the aromatic rings that cannot be described by the mentioned contributions. The base stacking can be rather reasonably approximated by simple molecular simulation methods based on well-calibrated common force fields although the force fields do not include nonadditivity of stacking, anisotropy of dispersion interactions, and some other effects. However, description of stacking association in condensed phase and understanding of the stacking role in biomolecules remain a difficult problem, as the net base stacking forces always act in a complex and context-specific environment. Moreover, the stacking forces are balanced with many other energy contributions. Differences in definition of stacking in experimental and theoretical studies are explained.
Electronic and vibrational optical activity of the set of neurohypophyseal hormones and their analogs was investigated to clarify the S-S bond solution conformation. The selected compounds include oxytocin (I), lysine vasopressin (II), arginine vasopressin (III), and their analogs (IV-IX), differing widely in their pharmacological properties. We have extended the already known electronic circular dichroism data by new information provided by vibrational circular dichroism (VCD) and Raman optical activity (ROA). The use of VCD brought additional details on three-dimensional structure of the chain reversal in the ring moiety and on its left handedness. Furthermore, Raman scattering and ROA allowed us to deduce the sense of the disulfide bond torsion.
- MeSH
- arginin vasopresin analogy a deriváty MeSH
- cirkulární dichroismus MeSH
- disulfidy chemie MeSH
- elektrony MeSH
- lypresin analogy a deriváty MeSH
- molekulární konformace MeSH
- optická otáčivost MeSH
- oxytocin analogy a deriváty MeSH
- Ramanova spektroskopie MeSH
- stereoizomerie MeSH
- torze mechanická MeSH
- vibrace MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
This work is a continuation of our effort to determine the structure responsible for expansion of the (CGG)(n) motif that results in fragile X chromosome syndrome. In our previous report, we demonstrated that the structure adopted by an oligonucleotide with this repeat sequence is not a quadruplex as was suggested by others. Here we demonstrate that (CGG) runs adopt another anomalous arrangement-a left-handed Z-DNA structure. The Z-DNA formation was induced by high salt and millimolar concentrations of Ni(2+) ions and likelihood of its formation increased with increasing number of repeats. In an oligonucleotide in which the CGG runs were interrupted by AGG triplets, as is observed in genomes of healthy individuals, the hairpin conformation was stabilized and Z-DNA formation was hindered. We show here that methylation of the (CGG) runs markedly stabilized Z-DNA formation. We hypothesize that rather than in the expansion process the Z-DNA may be formed by long, expanded (CGG) stretches that become hypermethylated; this would inhibit transcription resulting in disease.
- MeSH
- cirkulární dichroismus MeSH
- fragilní místa na chromozomu genetika MeSH
- konformace nukleové kyseliny MeSH
- lidé MeSH
- lidské chromozomy X genetika MeSH
- sekvence nukleotidů MeSH
- syndrom fragilního X genetika MeSH
- trinukleotidové repetice MeSH
- Z-DNA chemie genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
In an attempt to prepare a library of short oligoadenylate analogues featuring both the enzyme-stable internucleotide linkage and the 5'-O-methylphosphonate moiety and thus obtain a pool of potential RNase L agonists/antagonists, we studied the spontaneous polycondensation of the adenosin-5'-O-ylmethylphosphonic acid (p(c)A), an isopolar AMP analogue, and its imidazolide derivatives employing N,N'-dicyclohexylcarbodiimide under nonaqueous conditions and uranyl ions under aqueous conditions, respectively. The RP LC-MS analyses of the reaction mixtures per se, and those obtained after the periodate treatment, along with analyses and separations by capillary zone electrophoresis, allowed us to characterize major linear and cyclic oligoadenylates obtained. The structure of selected compounds was supported, after their isolation, by NMR spectroscopy. Ab initio calculation of the model structures simulating the AMP-imidazolide and p(c)A-imidazolide offered the explanation why the latter compound exerted, in contrast to AMP-imidazolide, a very low stability in aqueous solutions.
- MeSH
- adeninnukleotidy chemie metabolismus MeSH
- adenosinmonofosfát analogy a deriváty MeSH
- molekulární modely MeSH
- molekulární struktura MeSH
- nukleární magnetická rezonance biomolekulární MeSH
- oligoribonukleotidy chemie metabolismus MeSH
- oxidace-redukce MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Synthetic peptides corresponding to the sixth transmembrane segment (TMS6) of secondary-active transporter MntH (Proton-dependent Manganese Transporter) from Escherichia coli and its two mutations in the functionally important conserved histidine residue were used as a model for structure-function study of MntH. The secondary structure of the peptides was estimated in different environments using circular dichroism spectroscopy. These peptides interacted with and adopted helical conformations in lipid membranes. Electrophysiological experiments demonstrated that TMS6 was able to form multi-state ion channels in model biological membranes. Electrophysiological properties of these weakly cation-selective ion channels were strongly dependent on the surrounding pH. Manganese ion, as a physiological substrate of MntH, enhanced the conductivity of TMS6 channels, influenced the transition between closed and open states, and affected the peptide conformations. Moreover, functional properties of peptides carrying two different mutations of His(211) were analogous to in vivo functional characteristics of Nramp/MntH proteins mutated at homologous residues. Hence, a single functionally important TMS can retain some of the functional properties of the full-length protein. These findings could contribute to understanding the structure-function relationship at the molecular level. However it remains unclear to what extent the peptide-specific channel activity represents a functional aspect of the full-length membrane carrier protein.
- MeSH
- cirkulární dichroismus MeSH
- Escherichia coli genetika metabolismus MeSH
- iontové kanály chemie genetika metabolismus MeSH
- konformace proteinů MeSH
- liposomy MeSH
- mangan metabolismus MeSH
- metoda terčíkového zámku MeSH
- molekulární sekvence - údaje MeSH
- mutageneze cílená MeSH
- mutantní proteiny chemie genetika metabolismus MeSH
- peptidové fragmenty chemie genetika metabolismus MeSH
- proteiny přenášející kationty chemie genetika metabolismus MeSH
- proteiny z Escherichia coli chemie genetika metabolismus MeSH
- sekundární struktura proteinů MeSH
- sekvence aminokyselin MeSH
- substituce aminokyselin MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The interactions of valinomycin, macrocyclic depsipeptide antibiotic ionophore, with ammonium cation NH4+ have been investigated. Using quantum mechanical density functional theory (DFT) calculations, the most probable structure of the valinomycin-NH4+ complex species was predicted. In this complex, the ammonium cation is bound partly by three strong hydrogen bonds to three ester carbonyl oxygen atoms of valinomycin and partly by somewhat weaker hydrogen bonds to the remaining three ester carbonyl groups of the valinomycin ligand. The strength of the valinomycin-NH4+ complex was evaluated experimentally by capillary affinity electrophoresis. From the dependence of valinomycin effective electrophoretic mobility on the ammonium ion concentration in the background electrolyte, the apparent binding (association, stability) constant (Kb) of the valinomycin-NH4+ complex in methanol was evaluated as log Kb = 1.52 +/- 0.22.
