Muellner A.N.,Senckenberg Institute |
Muellner A.N.,Goethe University Frankfurt |
Schaefer H.,Senckenberg Institute |
Schaefer H.,Groningen Biomolecular science and Biotechnology Institute |
Lahaye R.,Services Geographiques
Molecular Ecology Resources | Year: 2011
There has been considerable debate regarding locus choice for DNA barcoding land plants. This is partly attributable to a shortage of comparable data from proposed candidate loci on a common set of samples. In this study, we evaluated main candidate plastid regions (rpoC1, rpoB, accD) and additional plastid markers (psbB, psbN, psbT exons and the trnS-trnG spacer) as well as the nuclear ribosomal spacer region (ITS1-5.8S-ITS2) in a group of land plants belonging to the mahogany family, Meliaceae. Across these samples, only ITS showed high levels of resolvability. Interspecific sharing of sequences from individual plastid loci was common. The combination of multiple loci did not improve performance. DNA barcoding with ITS alone revealed cryptic species and proved useful in identifying species listed in Convention on International Trade of Endangered Species appendixes. © 2011 Blackwell Publishing Ltd.
Risselada H.J.,Max Planck Institute for Biophysical Chemistry |
Marelli G.,University of Gottingen |
Fuhrmans M.,University of Gottingen |
Smirnova Y.G.,University of Gottingen |
And 3 more authors.
PLoS ONE | Year: 2012
Our molecular simulations reveal that wild-type influenza fusion peptides are able to stabilize a highly fusogenic pre-fusion structure, i.e. a peptide bundle formed by four or more trans-membrane arranged fusion peptides. We rationalize that the lipid rim around such bundle has a non-vanishing rim energy (line-tension), which is essential to (i) stabilize the initial contact point between the fusing bilayers, i.e. the stalk, and (ii) drive its subsequent evolution. Such line-tension controlled fusion event does not proceed along the hypothesized standard stalk-hemifusion pathway. In modeled influenza fusion, single point mutations in the influenza fusion peptide either completely inhibit fusion (mutants G1V and W14A) or, intriguingly, specifically arrest fusion at a hemifusion state (mutant G1S). Our simulations demonstrate that, within a line-tension controlled fusion mechanism, these known point mutations either completely inhibit fusion by impairing the peptide's ability to stabilize the required peptide bundle (G1V and W14A) or stabilize a persistent bundle that leads to a kinetically trapped hemifusion state (G1S). In addition, our results further suggest that the recently discovered leaky fusion mutant G13A, which is known to facilitate a pronounced leakage of the target membrane prior to lipid mixing, reduces the membrane integrity by forming a 'super' bundle. Our simulations offer a new interpretation for a number of experimentally observed features of the fusion reaction mediated by the prototypical fusion protein, influenza hemagglutinin, and might bring new insights into mechanisms of other viral fusion reactions. © 2012 Risselada et al.
Hansen D.F.,Kings College |
Neudecker P.,Kings College |
Vallurupalli P.,Kings College |
Vallurupalli P.,Indian Institute of Science |
And 2 more authors.
Journal of the American Chemical Society | Year: 2010
(Figure Presented) Fits of Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion profiles allow extraction of the kinetics and thermodynamics of exchange reactions that interconvert highly populated, ground state and low populated, excited state conformers. Structural information is also available in the form of chemical shift differences between the interconverting protein states. Here we present a very simple method for extracting x2 rotamer distributions of Leu side chains in 'invisible' excited protein states based on measurement of their 13Cδ1/ 13Cδ2 chemical shifts using methyl CPMG dispersion experiments. The methodology is applied to study the protein folding reaction of the Fyn SH3 domain. A uniform x2 rotamer distribution is obtained for Leu residues of the unfolded state, with each Leu occupying the trans and gauche+ conformations in a 2:1 ratio. By contrast, leucines of an 'invisible' Fyn SH3 domain folding intermediate show a much more heterogeneous distribution of x2 rotamer populations. The experiment provides an important tool toward the quantitative characterization of both the structural and dynamics properties of states that cannot be studied by other biophysical tools. © 2010 American Chemical Society.
Ramadurai S.,Groningen Biomolecular science and Biotechnology Institute |
Holt A.,Institute of Biomembranes |
Schafer L.V.,University of Groningen |
Krasnikov V.V.,Groningen Biomolecular science and Biotechnology Institute |
And 4 more authors.
Biophysical Journal | Year: 2010
We investigated the effect of amino acid composition and hydrophobic length of α-helical transmembrane peptides and the role of electrostatic interactions on the lateral diffusion of the peptides in lipid membranes. Model peptides of varying length and composition, and either tryptophans or lysines as flanking residues, were synthesized. The peptides were labeled with the fluorescent label Alexa Fluor 488 and incorporated into phospholipid bilayers of different hydrophobic thickness and composition. Giant unilamellar vesicles were formed by electroformation, and the lateral diffusion of the transmembrane peptides (and lipids) was determined by fluorescence correlation spectroscopy. In addition, we performed coarsegrained molecular-dynamics simulations of single peptides of different hydrophobic lengths embedded in planar membranes of different thicknesses. Both the experimental and simulation results indicate that lateral diffusion is sensitive to membrane thickness between the peptides and surrounding lipids. We did not observe a difference in the lateral diffusion of the peptides with respect to the presence of tryptophans or lysines as flanking residues. The specific lipid headgroup composition of the membrane has a much less pronounced impact on the diffusion of the peptides than does the hydrophobic thickness. © 2010 by the Biophysical Society.
Wijma H.J.,Groningen Biomolecular science and Biotechnology Institute |
Janssen D.B.,Groningen Biomolecular science and Biotechnology Institute
FEBS Journal | Year: 2013
Computational protein design is becoming a powerful tool for tailoring enzymes for specific biotechnological applications. When applied to existing enzymes, computational re-design makes it possible to obtain orders of magnitude improvement in catalytic activity towards a new target substrate. Computational methods also allow the design of completely new active sites that catalyze reactions that are not known to occur in biological systems. If initial designs display modest catalytic activity, which is often the case, this may be improved by iterative cycles of computational design or by follow-up engineering through directed evolution. Compared to established protein engineering methods such as directed evolution and structure-based mutagenesis, computational design allows for much larger jumps in sequence space; for example, by introducing more than a dozen mutations in a single step or by introducing loops that provide new functional interactions. Recent advances in the computational design toolbox, which include new backbone re-design methods and the use of molecular dynamics simulations to better predict the catalytic activity of designed variants, will further enhance the use of computational tools in enzyme engineering. Computational protein design is becoming a powerful tool for tailoring enzymes for specific biotechnological applications. Compared to established protein engineering methods, computational design allows much larger jumps in sequence space. Recent advances in the computational design toolbox, which include new backbone redesign methods and the use of molecular dynamics simulations, will increase the possibilities for computational enzyme engineering. © 2013 FEBS.