@article{8482, abstract = {The SOFAST-HMQC experiment [P. Schanda, B. Brutscher, Very fast two-dimensional NMR spectroscopy for real-time investigation of dynamic events in proteins on the time scale of seconds, J. Am. Chem. Soc. 127 (2005) 8014–8015] allows recording two-dimensional correlation spectra of macromolecules such as proteins in only a few seconds acquisition time. To achieve the highest possible sensitivity, SOFAST-HMQC experiments are preferably performed on high-field NMR spectrometers equipped with cryogenically cooled probes. The duty cycle of over 80% in fast-pulsing SOFAST-HMQC experiments, however, may cause problems when using a cryogenic probe. Here we introduce SE-IPAP-SOFAST-HMQC, a new pulse sequence that provides comparable sensitivity to standard SOFAST-HMQC, while avoiding heteronuclear decoupling during 1H detection, and thus significantly reducing the radiofrequency load of the probe during the experiment. The experiment is also attractive for fast and sensitive measurement of heteronuclear one-bond spin coupling constants.}, author = {Kern, Thomas and Schanda, Paul and Brutscher, Bernhard}, issn = {1090-7807}, journal = {Journal of Magnetic Resonance}, keywords = {Nuclear and High Energy Physics, Biophysics, Biochemistry, Condensed Matter Physics}, number = {2}, pages = {333--338}, publisher = {Elsevier}, title = {{Sensitivity-enhanced IPAP-SOFAST-HMQC for fast-pulsing 2D NMR with reduced radiofrequency load}}, doi = {10.1016/j.jmr.2007.11.015}, volume = {190}, year = {2008}, } @article{11116, abstract = {The metazoan nuclear envelope (NE) breaks down and re-forms during each cell cycle. Nuclear pore complexes (NPCs), which allow nucleocytoplasmic transport during interphase, assemble into the re-forming NE at the end of mitosis. Using in vitro NE assembly, we show that the vertebrate homologue of MEL-28 (maternal effect lethal), a recently discovered NE component in Caenorhabditis elegans, functions in postmitotic NPC assembly. MEL-28 interacts with the Nup107–160 complex (Nup for nucleoporin), an important building block of the NPC, and is essential for the recruitment of the Nup107–160 complex to chromatin. We suggest that MEL-28 acts as a seeding point for NPC assembly.}, author = {Franz, Cerstin and Walczak, Rudolf and Yavuz, Sevil and Santarella, Rachel and Gentzel, Marc and Askjaer, Peter and Galy, Vincent and HETZER, Martin W and Mattaj, Iain W and Antonin, Wolfram}, issn = {1469-3178}, journal = {EMBO reports}, keywords = {Genetics, Molecular Biology, Biochemistry}, number = {2}, pages = {165--172}, publisher = {EMBO}, title = {{MEL‐28/ELYS is required for the recruitment of nucleoporins to chromatin and postmitotic nuclear pore complex assembly}}, doi = {10.1038/sj.embor.7400889}, volume = {8}, year = {2007}, } @article{8485, abstract = {High signal to noise is a necessity for the quantification of NMR spectral parameters to be translated into accurate and precise restraints on protein structure and dynamics. An important source of long-range structural information is obtained from 1H–1H residual dipolar couplings (RDCs) measured for weakly aligned molecules. For sensitivity reasons, such measurements are generally performed on highly deuterated protein samples. Here we show that high sensitivity is also obtained for protonated protein samples if the pulse schemes are optimized in terms of longitudinal relaxation efficiency and J-mismatch compensated coherence transfer. The new sensitivity-optimized quantitative J-correlation experiment yields important signal gains reaching factors of 1.5 to 8 for individual correlation peaks when compared to previously proposed pulse schemes.}, author = {Schanda, Paul and Lescop, Ewen and Falge, Mirjam and Sounier, Rémy and Boisbouvier, Jérôme and Brutscher, Bernhard}, issn = {0925-2738}, journal = {Journal of Biomolecular NMR}, keywords = {Spectroscopy, Biochemistry}, pages = {47--55}, publisher = {Springer Nature}, title = {{Sensitivity-optimized experiment for the measurement of residual dipolar couplings between amide protons}}, doi = {10.1007/s10858-006-9138-2}, volume = {38}, year = {2007}, } @article{8486, abstract = {A technique is described that allows reducing acquisition times of multidimensional NMR experiments by extensive spectral folding. The method is simple and has many interesting applications for NMR studies of molecular structure, dynamics, and kinetics.}, author = {Lescop, Ewen and Schanda, Paul and Rasia, Rodolfo and Brutscher, Bernhard}, issn = {0002-7863}, journal = {Journal of the American Chemical Society}, keywords = {Colloid and Surface Chemistry, Biochemistry, General Chemistry, Catalysis}, number = {10}, pages = {2756--2757}, publisher = {American Chemical Society}, title = {{Automated spectral compression for fast multidimensional NMR and increased time resolution in real-time NMR spectroscopy}}, doi = {10.1021/ja068949u}, volume = {129}, year = {2007}, } @article{8487, abstract = {Following unidirectional biophysical events such as the folding of proteins or the equilibration of binding interactions, requires experimental methods that yield information at both atomic-level resolution and at high repetition rates. Toward this end a number of different approaches enabling the rapid acquisition of 2D NMR spectra have been recently introduced, including spatially encoded “ultrafast” 2D NMR spectroscopy and SOFAST HMQC NMR. Whereas the former accelerates acquisitions by reducing the number of scans that are necessary for completing arbitrary 2D NMR experiments, the latter operates by reducing the delay between consecutive scans while preserving sensitivity. Given the complementarities between these two approaches it seems natural to combine them into a single tool, enabling the acquisition of full 2D protein NMR spectra at high repetition rates. We demonstrate here this capability with the introduction of “ultraSOFAST” HMQC NMR, a spatially encoded and relaxation-optimized approach that can provide 2D protein correlation spectra at ∼1 s repetition rates for samples in the ∼2 mM concentration range. The principles, relative advantages, and current limitations of this new approach are discussed, and its application is exemplified with a study of the fast hydrogen−deuterium exchange characterizing amide sites in Ubiquitin.}, author = {Gal, Maayan and Schanda, Paul and Brutscher, Bernhard and Frydman, Lucio}, issn = {0002-7863}, journal = {Journal of the American Chemical Society}, keywords = {Colloid and Surface Chemistry, Biochemistry, General Chemistry, Catalysis}, number = {5}, pages = {1372--1377}, publisher = {American Chemical Society}, title = {{UltraSOFAST HMQC NMR and the repetitive acquisition of 2D protein spectra at Hz rates}}, doi = {10.1021/ja066915g}, volume = {129}, year = {2007}, } @article{8488, abstract = {We demonstrate for different protein samples that three-dimensional HNCO and HNCA correlation spectra may be recorded in a few minutes acquisition time using the band-selective excitation short-transient sequences presented here. This opens new perspectives for the NMR structural investigation of unstable protein samples and real-time site-resolved studies of protein kinetics.}, author = {Schanda, Paul and Van Melckebeke, Hélène and Brutscher, Bernhard}, issn = {0002-7863}, journal = {Journal of the American Chemical Society}, keywords = {Colloid and Surface Chemistry, Biochemistry, General Chemistry, Catalysis}, number = {28}, pages = {9042--9043}, publisher = {American Chemical Society}, title = {{Speeding up three-dimensional protein NMR experiments to a few minutes}}, doi = {10.1021/ja062025p}, volume = {128}, year = {2006}, } @article{8490, abstract = {We demonstrate the feasibility of recording 1H–15N correlation spectra of proteins in only one second of acquisition time. The experiment combines recently proposed SOFAST-HMQC with Hadamard-type 15N frequency encoding. This allows site-resolved real-time NMR studies of kinetic processes in proteins with an increased time resolution. The sensitivity of the experiment is sufficient to be applicable to a wide range of molecular systems available at millimolar concentration on a high magnetic field spectrometer.}, author = {Schanda, Paul and Brutscher, Bernhard}, issn = {1090-7807}, journal = {Journal of Magnetic Resonance}, keywords = {Nuclear and High Energy Physics, Biophysics, Biochemistry, Condensed Matter Physics}, number = {2}, pages = {334--339}, publisher = {Elsevier}, title = {{Hadamard frequency-encoded SOFAST-HMQC for ultrafast two-dimensional protein NMR}}, doi = {10.1016/j.jmr.2005.10.007}, volume = {178}, year = {2006}, } @article{13428, abstract = {Mixtures of oppositely charged nanoparticles of various sizes and charge ratios precipitate only at the point of electroneutrality. This phenomenonspecific to the nanoscale and reminiscent of threshold precipitation of ionsis a consequence of the formation of core-and-shell nanoparticle aggregates, in which the shells are composed of like-charged particles and are stabilized by efficient electrostatic screening.}, author = {Kalsin, Alexander M. and Kowalczyk, Bartlomiej and Smoukov, Stoyan K. and Klajn, Rafal and Grzybowski, Bartosz A.}, issn = {1520-5126}, journal = {Journal of the American Chemical Society}, keywords = {Colloid and Surface Chemistry, Biochemistry, General Chemistry, Catalysis}, number = {47}, pages = {15046--15047}, publisher = {American Chemical Society}, title = {{Ionic-like behavior of oppositely charged nanoparticles}}, doi = {10.1021/ja0642966}, volume = {128}, year = {2006}, } @article{8491, abstract = {Fast multidimensional NMR with a time resolution of a few seconds provides a new tool for high throughput screening and site-resolved real-time studies of kinetic molecular processes by NMR. Recently we have demonstrated the feasibility to record protein 1H–15N correlation spectra in a few seconds of acquisition time using a new SOFAST-HMQC experiment (Schanda and Brutscher (2005) J. Am. Chem. Soc. 127, 8014). Here, we investigate in detail the performance of SOFAST-HMQC to record 1H–15N and 1H−13C correlation spectra of proteins of different size and at different magnetic field strengths. Compared to standard 1H–15N correlation experiments SOFAST-HMQC provides a significant gain in sensitivity, especially for fast repetition rates. Guidelines are provided on how to set up SOFAST-HMQC experiments for a given protein sample. In addition, an alternative pulse scheme, IPAP-SOFAST-HMQC is presented that allows application on NMR spectrometers equipped with cryogenic probes, and fast measurement of one-bond 1H–13C and 1H–15N scalar and residual dipolar coupling constants.}, author = {Schanda, Paul and Kupče, Ēriks and Brutscher, Bernhard}, issn = {0925-2738}, journal = {Journal of Biomolecular NMR}, keywords = {Spectroscopy, Biochemistry}, number = {4}, pages = {199--211}, publisher = {Springer Nature}, title = {{SOFAST-HMQC experiments for recording two-dimensional deteronuclear correlation spectra of proteins within a few seconds}}, doi = {10.1007/s10858-005-4425-x}, volume = {33}, year = {2005}, } @article{8492, abstract = {We demonstrate for different protein samples that 2D 1H−15N correlation NMR spectra can be recorded in a few seconds of acquisition time using a new band-selective optimized flip-angle short-transient heteronuclear multiple quantum coherence experiment. This has enabled us to measure fast hydrogen−deuterium exchange rate constants along the backbone of a small globular protein fragment by real-time 2D NMR.}, author = {Schanda, Paul and Brutscher, Bernhard}, issn = {0002-7863}, journal = {Journal of the American Chemical Society}, keywords = {Colloid and Surface Chemistry, Biochemistry, General Chemistry, Catalysis}, number = {22}, pages = {8014--8015}, publisher = {American Chemical Society}, title = {{Very fast two-dimensional NMR spectroscopy for real-time investigation of dynamic events in proteins on the time scale of seconds}}, doi = {10.1021/ja051306e}, volume = {127}, year = {2005}, } @article{12203, abstract = {Geranylgeranyl diphosphate synthase (GGPPS, EC: 2.5.1.29) catalyzes the biosynthesis of geranylgeranyl diphosphate (GGPP), which is a key precursor for ginkgolide biosynthesis. Here we reported for the first time the cloning of a new full-length cDNA encoding GGPPS from the living fossil plant Ginkgo biloba. The full-length cDNA encoding G. biloba GGPPS (designated as GbGGPPS) was 1657bp long and contained a 1176bp open reading frame encoding a 391 amino acid protein. Comparative analysis showed that GbGGPPS possessed a 79 amino acid transit peptide at its N-terminal, which directed GbGGPPS to target to the plastids. Bioinformatic analysis revealed that GbGGPPS was a member of polyprenyltransferases with two highly conserved aspartate-rich motifs like other plant GGPPSs. Phylogenetic tree analysis indicated that plant GGPPSs could be classified into two groups, angiosperm and gymnosperm GGPPSs, while GbGGPPS had closer relationship with gymnosperm plant GGPPSs.}, author = {Liao, Zhihua and Chen, Min and Gong, Yifu and Guo, Liang and Tan, Qiumin and Feng, Xiaoqi and Sun, Xiaofen and Tan, Feng and Tang, Kexuan}, issn = {1042-5179}, journal = {DNA Sequence}, keywords = {Endocrinology, Genetics, Molecular Biology, Biochemistry}, number = {2}, pages = {153--158}, publisher = {Informa UK Limited}, title = {{A new geranylgeranyl Diphosphate synthase gene from Ginkgo biloba, which intermediates the biosynthesis of the key precursor for ginkgolides}}, doi = {10.1080/10425170410001667348}, volume = {15}, year = {2004}, } @article{11122, abstract = {Nuclear pore complexes (NPCs) are large multiprotein assemblies that allow traffic between the cytoplasm and the nucleus. During mitosis in higher eukaryotes, the Nuclear Envelope (NE) breaks down and NPCs disassemble. How NPCs reassemble and incorporate into the NE upon mitotic exit is poorly understood. We demonstrate a function for the conserved Nup107-160 complex in this process. Partial in vivo depletion of Nup133 or Nup107 via RNAi in HeLa cells resulted in reduced levels of multiple nucleoporins and decreased NPC density in the NE. Immunodepletion of the entire Nup107-160 complex from in vitro nuclear assembly reactions produced nuclei with a continuous NE but no NPCs. This phenotype was reversible only if Nup107-160 complex was readded before closed NE formation. Depletion also prevented association of FG-repeat nucleoporins with chromatin. We propose a stepwise model in which postmitotic NPC assembly initiates on chromatin via early recruitment of the Nup107-160 complex.}, author = {Walther, Tobias C. and Alves, Annabelle and Pickersgill, Helen and Loı̈odice, Isabelle and HETZER, Martin W and Galy, Vincent and Hülsmann, Bastian B. and Köcher, Thomas and Wilm, Matthias and Allen, Terry and Mattaj, Iain W. and Doye, Valérie}, issn = {0092-8674}, journal = {Cell}, keywords = {General Biochemistry, Genetics and Molecular Biology}, number = {2}, pages = {195--206}, publisher = {Elsevier}, title = {{The conserved Nup107-160 complex is critical for nuclear pore complex assembly}}, doi = {10.1016/s0092-8674(03)00235-6}, volume = {113}, year = {2003}, } @article{13436, abstract = {Cross-metathesis reactions of α,β-unsaturated sulfones and sulfoxides in the presence of molybdenum and ruthenium pre-catalysts were tested. A selective metahesis reaction was achieved between functionalized terminal olefins and vinyl sulfones by using the ‘second generation’ ruthenium catalysts 1c–h while the highly active Schrock catalyst 1b was found to be functional group incompatible with vinyl sulfones. The cross-metathesis products were isolated in good yields with an excellent (E)-selectivity. Both the molybdenum and ruthenium-based complexes were, however, incompatible with α,β- and β,γ-unsaturated sulfoxides.}, author = {Michrowska, Anna and Bieniek, Michał and Kim, Mikhail and Klajn, Rafal and Grela, Karol}, issn = {1464-5416}, journal = {Tetrahedron}, keywords = {Organic Chemistry, Drug Discovery, Biochemistry}, number = {25}, pages = {4525--4531}, publisher = {Elsevier}, title = {{Cross-metathesis reaction of vinyl sulfones and sulfoxides}}, doi = {10.1016/s0040-4020(03)00682-3}, volume = {59}, year = {2003}, } @article{11124, abstract = {Ran GTPase plays important roles in nucleocytoplasmic transport in interphase [1, 2] and in both spindle formation and nuclear envelope (NE) assembly during mitosis [3, 4, 5]. The latter functions rely on the presence of high local concentrations of GTP-bound Ran near mitotic chromatin [3, 4, 5]. RanGTP localization has been proposed to result from the association of Ran's GDP/GTP exchange factor, RCC1, with chromatin [6, 7, 8, 9], but Ran is shown here to bind directly to chromatin in two modes, either dependent or independent of RCC1, and, where bound, to increase the affinity of chromatin for NE membranes. We propose that the Ran binding capacity of chromatin contributes to localized spindle and NE assembly.}, author = {Bilbao-Cortés, Daniel and HETZER, Martin W and Längst, Gernot and Becker, Peter B. and Mattaj, Iain W.}, issn = {0960-9822}, journal = {Current Biology}, keywords = {General Agricultural and Biological Sciences, General Biochemistry, Genetics and Molecular Biology}, number = {13}, pages = {1151--1156}, publisher = {Elsevier BV}, title = {{Ran binds to chromatin by two distinct mechanisms}}, doi = {10.1016/s0960-9822(02)00927-2}, volume = {12}, year = {2002}, } @article{13438, abstract = {ICln is an ion channel identified by expression cloning using a cDNA library from Madin-Darby canine kidney cells. In all organisms tested so far, only one transcript for the ICln protein could be identified. Here we show that two splice variants of the ICln ion channel can be found in Caenorhabditis elegans. Moreover, we show that these two splice variants of the ICln channel protein, which we termed IClnN1 and IClnN2, can be functionally reconstituted and tested in an artificial lipid bilayer. In these experiments, the IClnN1-induced currents showed no voltage-dependent inactivation, whereas the IClnN2-induced currents fully inactivated at positive potentials. The molecular entity responsible for the voltage-dependent inactivation of IClnN2 is a cluster of positively charged amino acids encoded by exon 2a, which is absent in IClnN1. Our experiments suggest a mechanism of channel inactivation that is similar to the “ball and chain” model proposed for the Shaker potassium channel,i.e. a cluster of positively charged amino acids hinders ion permeation through the channel by a molecular and voltage-dependent interaction at the inner vestibulum of the pore. This hypothesis is supported by the finding that synthetic peptides with the same amino acid sequence as the positive cluster can transform the IClnN1-induced current to the current observed after reconstitution of IClnN2. Furthermore, we show that the nematode ICln gene is embedded in an operon harboring two additional genes, which we termed Nx and Ny. Co-reconstitution of Nx and IClnN2 and functional analysis of the related currents revealed a functional interaction between the two proteins, as evidenced by the fact that the IClnN2-induced current in the presence of Nx was no longer voltage-sensitive. The experiments described indicate that the genome organization in nematodes allows an effective approach for the identification of functional partner proteins of ion channels.}, author = {Fürst, Johannes and Ritter, Markus and Rudzki, Jakob and Danzl, Johann G and Gschwentner, Martin and Scandella, Elke and Jakab, Martin and König, Matthias and Oehl, Bernhard and Lang, Florian and Deetjen, Peter and Paulmichl, Markus}, issn = {0021-9258}, journal = {Journal of Biological Chemistry}, keywords = {Cell Biology, Molecular Biology, Biochemistry}, number = {6}, pages = {4435--4445}, publisher = {Elsevier}, title = {{ICln Ion channel splice variants in Caenorhabditis elegans}}, doi = {10.1074/jbc.m107372200}, volume = {277}, year = {2002}, }