Molecular dynamics (MD) simulations have been used to characterize the effects of backbone N-amination of residues in a model β-hairpin peptide. This modification is of considerable interest as ...N-aminated peptides have been shown to inhibit amyloid-type aggregation. Six derivatives of the β-hairpin peptide, which contain one, two, or four N-aminated residues, have been studied. For each peptide 100 ns MD simulations starting from the folded β-hairpin structure were performed. The effects of the N-amination prove to be very sequence dependent. N-Amination of a residue involved in interstrand hydrogen bonding (Val3) leads to unfolding of the β-hairpin, whereas N-amination of a residue toward the C-terminus (Leu11) gives fraying at the termini of the peptide. In the other derivatives the peptide remains folded, with increasing levels of N-amination reducing the right-handed twist of the β-hairpin and favoring population of a type II′ rather than a type I′ β-turn. MD simulations (100 ns) have also been run for each peptide starting from an unfolded extended chain. Here, the peptide with four N-aminated residues shows the most folding into the β-hairpin (34%). Analysis of the simulations shows that N-amination favors the population of β (φ, ψ) conformations by the preceding residue due to, at least in part, a network of weak NH2(i)–CO(i) and NH2(i)–CO(i–2) hydrogen bonds. It also leads to a reduction of misfolding because of changes in the hydrogen-bonding potential. Both of these features help funnel the peptide to the folded β-hairpin structure. The conformational insights provided through this work give a firm foundation for the design of N-aminated peptide inhibitors for modulating protein–protein interactions and aggregation.
Diatoms of the genus
H.Peragallo are known to produce domoic acid (DA), a toxin involved in amnesic shellfish poisoning (ASP). Strains of the same species are often classified as both toxic and ...nontoxic, and it is largely unknown whether this difference is also genetic. In the Northern Adriatic Sea, there are virtually no cases of ASP, but DA occasionally occurs in shellfish samples. So far, three species-
(Cleve) Heiden
(H. Takano) H. Takano, and
Lundholm, Moestrup, & Hasle-have been identified as producers of DA in the Adriatic Sea. By means of enzme-linked immunosorbent assay (ELISA), high-performance liquid chromatography with UV and visible spectrum detection (HPLC-UV/VIS), and liquid chromatography with tandem mass spectrometry (LC-MS/MS), we reconfirmed the presence of DA in
and
and detect for the first time in the Adriatic Sea DA in
Lundholm, & Moestrup. Furthermore, we attempted to answer the question of the distribution of DA production among
species and strains by sequencing the internal transcribed spacer (ITS) phylogenetic marker and the
DA biosynthesis gene and coupling this with toxicity data. Results show that all subclades of the
genus contain toxic species and that toxicity appears to be strain dependent, often with geographic partitioning. Amplification of
was successful only in toxic strains of
and the presence of the genetic architecture for DA production in non-toxic strains was thus not confirmed.
Computer-based molecular simulation techniques are increasingly used to interpret experimental data on biomolecular systems at an atomic level. Direct comparison between experiment and simulation is, ...however, seldom straightforward. The available experimental data are limited in scope and generally correspond to averages over both time and space. A critical analysis of the various factors that may influence the apparent degree of agreement between the results of simulations and experimentally measured quantities is presented and illustrated using examples from recent literature.
GROMOS++ is a set of C++ programs for pre- and postprocessing of molecular dynamics simulation trajectories and as such is part of the GROningen MOlecular Simulation software for (bio)molecular ...simulation. It contains more than 70 programs that can be used to prepare data for the production of molecular simulation trajectories and to analyze these. These programs are reviewed and the various structural, dynamic, and thermodynamic quantities that can be analyzed using time series, correlation functions, and distributions are described together with technical aspects of their implementation in GROMOS. A few examples of the use of GROMOS++ for the analysis of MD trajectories are given. A full list of all GROMOS++ programs, together with an indication of their capabilities, is given in the Appendix.
Epothilones are among the most potent chemotherapeutic drugs used for the treatment of cancer. Epothilone A (EpoA), a natural product, is a macrocyclic molecule containing 34 non-hydrogen atoms and a ...thiazole side chain. NMR studies of EpoA in aqueous solution, unbound as well as bound to αβ-tubulin, and unbound in dimethyl sulfoxide (DMSO) solution have delivered sets of nuclear Overhauser effect (NOE) atom–atom distance bounds, but no structures based on NMR data are present in structural data banks. X-ray diffraction of crystals has provided structures of EpoA unbound and bound to αβ-tubulin. Since both crystal structures derived from X-ray diffraction intensities do not completely satisfy the three available sets of NOE distance bounds for EpoA, molecular dynamics (MD) simulations have been employed to obtain conformational ensembles in aqueous and in DMSO solution that are compatible with the respective NOE data. It was found that EpoA displays a larger conformational variability in DMSO than in water and the two conformational ensembles show little overlap. Yet, they both provide conformational scaffolds that are energetically accessible at physiological temperature and pressure.
Background
Undiagnosed low-grade prosthetic joint infections (PJI) are recognized as an important reason for early failure of presumably aseptic revisions. Preoperatively administered antimicrobial ...prophylaxis reduces the incidence of PJI but it may reduce the sensitivity of microbiologic periprosthetic tissue cultures and consequently increase the incidence of undiagnosed septic prosthetic joint failures, which can lead to catastrophic serial revisions.
Questions/purposes
We wished to determine whether administration of preoperative antibiotics decreases the likelihood of diagnosing PJI in patients undergoing revision hip or knee arthroplasty in whom infection is suspected.
Methods
We prospectively enrolled and evaluated 40 patients (29 with THAs and 11 with TKAs) who met the following inclusion criteria: older than 18 years, with suspected PJI of unknown cause, undergoing surgical revision. After arthrotomy, three tissue samples were obtained for microbiologic analysis and diagnosis, and antimicrobial prophylaxis (cefazolin 2 g intravenously) then was administered. Later during the procedure, but before débridement and irrigation, the second set of three tissue samples was obtained from the same surgical area and was cultured. Tissue concentration of prophylactic antibiotic was verified with the second set of samples. A positive culture result was defined as one or more positive cultures (growth on agar at or before 14 days). We then compared the yield on the microbiologic cultures obtained before administration of antibiotics with the yield on the cultures obtained after antibiotics were administered. An a priori analysis was performed; with the numbers available, we had 98% power to detect a difference in diagnostic sensitivity of 33%.
Results
With the numbers available, we found no difference in the likelihood that an infection would be diagnosed between the samples obtained before and after administration of antimicrobial prophylaxis (odds ratio OR for positive microbial culture = 0.99; 95% CI, 0.40–2.48; p = 0.99). All measured tissue concentrations of cefazolin were greater than the minimum inhibitory concentration, therefore we found that antibiotic prophylaxis was adequate at the time of second-set tissue specimen recovery.
Conclusions
Results from this small, prospective series suggest that preoperative antimicrobial prophylaxis may be administered safely even in patients undergoing revision hip or knee arthroplasty in which microbiologic sampling is planned without compromising the diagnostic sensitivity of tissue sample cultures. However, before applying our results more generally, our findings need to be confirmed in larger, multicenter studies that would allow evaluation by sex, procedure, bacteriology, and other potentially important factors.
Level of Evidence
Level I, diagnostic study.
With the aim to gain a better understanding of the various driving forces that govern sequence specific DNA minor groove binding, we performed a thermodynamic analysis of netropsin binding to an ...AT-containing and to a set of six mixed AT/GC-containing binding sequences in the DNA minor groove. The relative binding free energies obtained using molecular dynamics simulations and free energy calculations show significant variations with the binding sequence. While the introduction of a GC base pair in the middle or close to the middle of the binding site is unfavorable for netropsin binding, a GC base pair at the end of the binding site appears to have no negative influence on the binding. The results of the structural and energetic analyses of the netropsin−DNA complexes reveal that the differences in the calculated binding affinities cannot be explained solely in terms of netropsin−DNA hydrogen-bonding or interaction energies. In addition, solvation effects and entropic contributions to the relative binding free energy provide a more complete picture of the various factors determining binding. Analysis of the relative binding entropy indicates that its magnitude is highly sequence-dependent, with the ratio |TΔΔS|/|ΔΔH| ranging from 0.07 for the AAAGA to 1.7 for the AAGAG binding sequence, respectively.
The cyclic octa-peptide octreotide and its derivatives are used as diagnostics and therapeutics in relation to particular types of cancers. This led to investigations of their conformational ...properties using spectroscopic, NMR and CD, methods. A CF
-substituted derivative, that was designed to stabilize the dominant octreotide conformer responsible for receptor binding, turned out to have a lower affinity. The obtained spectroscopic data were interpreted as to show an increased flexibility of the CF
derivative compared to the unsubstituted octreotide, which could then explain the lower affinity. In this article, we use MD simulation without and with time-averaged NOE distance and time-averaged local-elevation
J-coupling restraining representing experimental NMR data to determine the conformational properties of the different peptides in the different solvents for which experimental data are available, that are compatible with the NOE atom-atom distance bounds and the
J
-couplings as derived from the NMR measurements. The conformational ensembles show that the CF
substitution in combination with the change of solvent from water to methanol leads to a decrease in flexibility and a shift in the populations of the dominant conformers that are compatible with the experimental data.
During the past half century, the number and accuracy of experimental techniques that can deliver values of observables for biomolecular systems have been steadily increasing. The conversion of a ...measured value Qexp of an observable quantity Q into structural information is, however, a task beset with theoretical and practical problems: 1) insufficient or inaccurate values of Qexp, 2) inaccuracies in the function Q(r→)
used to relate the quantity Q to structure r→
, 3) how to account for the averaging inherent in the measurement of Qexp, 4) how to handle the possible multiple‐valuedness of the inverse r→(Q)
of the function Q(r→)
, to mention a few. These apply to a variety of observable quantities Q and measurement techniques such as X‐ray and neutron diffraction, small‐angle and wide‐angle X‐ray scattering, free‐electron laser imaging, cryo‐electron microscopy, nuclear magnetic resonance, electron paramagnetic resonance, infrared and Raman spectroscopy, circular dichroism, Förster resonance energy transfer, atomic force microscopy and ion‐mobility mass spectrometry. The process of deriving structural information from measured data is reviewed with an eye to non‐experts and newcomers in the field using examples from the literature of the effect of the various choices and approximations involved in the process. A list of choices to be avoided is provided.
Making the right choice: Deriving structural information from experimentally measured data is a process beset with theoretical and practical problems. The effects of the various assumptions and approximations involved in the process of biomolecular structure determination are discussed and a list of choices to be avoided is provided.
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