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151.
P Burkhard P Dominici C Borri-Voltattorni J N Jansonius V N Malashkevich 《Nature structural biology》2001,8(11):963-967
DOPA decarboxylase (DDC) is responsible for the synthesis of the key neurotransmitters dopamine and serotonin via decarboxylation of L-3,4-dihydroxyphenylalanine (L-DOPA) and L-5-hydroxytryptophan, respectively. DDC has been implicated in a number of clinic disorders, including Parkinson's disease and hypertension. Peripheral inhibitors of DDC are currently used to treat these diseases. We present the crystal structures of ligand-free DDC and its complex with the anti-Parkinson drug carbiDOPA. The inhibitor is bound to the enzyme by forming a hydrazone linkage with the cofactor, and its catechol ring is deeply buried in the active site cleft. The structures provide the molecular basis for the development of new inhibitors of DDC with better pharmacological characteristics. 相似文献
152.
The prasinophyte genera Scherffelia and Tetraselmis are the only genera that form a cell wall by an extracellular fusion of scales called a theca. We established a protocol for the production of protoplasts from Scherffelia dubia Pascher emend. Melkonian et Preisig using 3 mM Ellman's reagent (5,5′‐dithio‐bis‐2‐nitrobenozoic acid [DTNB]). Protoplasts analyzed by EM lacked flagella and thecae but were otherwise similar to control cells. In response to treatment with DTNB, many protoplasts synthesized new thecal scales in the Golgi apparatus, indicating that cells attempted to regenerate new cell walls. However, complete regeneration of the thecae only occurred once DTNB was washed out from the medium. At higher DTNB concentrations (5 mM), two protoplasts were found within the parental cell wall and scales accumulated between the plasma membrane of the protoplasts and the original theca but failed to form a new theca. 相似文献
153.
The thecate green flagellate Scherffelia dubia (Perty) Pascher divides within the parental cell wall into two progeny cells. It sheds all four flagella before cell division, and the maturing progeny cells regenerate new walls and flagella. By synchronizing cell division, we observed mitosis, cytokinesis, cell maturation, flagella extension, and cell wall formation via differential interference contrast microscopy of live cells and serial thin‐section EM. Synthesis of thecal and flagellar scales is spatially and temporally strictly separated. Flagellar scales are collected in a pool during late interphase. Before prophase, Golgi stacks divide, flagella are shed, the parental theca separates from the plasma membrane, and flagellar scales are deposited on the plasma membrane near the flagellar bases. At prophase, Golgi bodies start to synthesize thecal scales, continuing into interphase after cytokinesis. During cytokinesis, vesicles containing thecal scales coalesce near the cell posterior, forming a cleavage furrow that is initially oriented slightly diagonal to the longitudinal cell axis but later becomes transverse. After the progeny nuclei have moved into opposite directions, resulting in a “head to tail” orientation of the progeny cells, theca biogenesis is completed and flagellar scale synthesis resumes. Progeny cells emerge through a hole near the posterior end of the parental theca with four flagella of about 8 μm long. The precise timing of flagellar and thecal scale synthesis appears to be an evolutionary adaptation in a scaly green flagellate for the thecal condition, necessary for the evolution of the phycoplast and thus multicellularity in the Chlorophyta. 相似文献
154.
Johannes Oehlke Gerd Wallukat Yvonne Wolf Angelika Ehrlich Burkhard Wiesner Hartmut Berger Michael Bienert 《European journal of biochemistry》2004,271(14):3043-3049
In order to evaluate the ability of the cell-penetrating alpha-helical amphipathic model peptide KLALKLALKALKAALKLA-NH(2) (MAP) to deliver peptide nucleic acids (PNAs) into mammalian cells, MAP was covalently linked to the 12-mer PNA 5'-GGAGCAGGAAAG-3' directed against the mRNA of the nociceptin/orphanin FQ receptor. The cellular uptake of both the naked PNA and its MAP-conjugate was studied by means of capillary electrophoresis combined with laser-induced fluorescence detection, confocal laser scanning microscopy and fluorescence-activated cell sorting. Incubation with the fluorescein-labelled PNA-peptide conjugate led to three- and eightfold higher intracellular concentrations in neonatal rat cardiomyocytes and CHO cells, respectively, than found after exposure of the cells to the naked PNA. Correspondingly, pretreatment of spontaneously-beating neonatal rat cardiomyocytes with the PNA-peptide conjugate and the naked PNA slowed down the positive chronotropic effect elicited by the neuropeptide nociceptin by 10- and twofold, respectively. The main reasons for the higher bioavailability of the PNA-peptide conjugate were found to be a more rapid cellular uptake in combination with a lowered re-export and resistance against influences of serum. 相似文献
155.
Burkhard Luy Alexander Diener Rolf-Peter Hummel Ernst Sturm Wolf-Rüdiger Ulrich Christian Griesinger 《European journal of biochemistry》2004,271(11):2076-2085
The solution structure of a recombinant mutant [rSP-C (FFI)] of the human surfactant-associated protein C (hSP-C) in a mixture of chloroform and methanol was determined by high-resolution NMR spectroscopy. rSP-C (FFI) contains a helix from Phe5 to the C-terminal Leu34 and is thus longer by two residues than the helix of porcine SP-C (pSP-C), which is reported to start at Val7 in the same solvent. Two sets of resonances at the C-terminus of the peptide were observed, which are explained by low-order oligomerization, probably dimerization of rSP-C (FFI) in its alpha-helical form. The dimerization may be induced by hydrogen bonding of the C-terminal carboxylic groups or by the strictly conserved C-terminal heptapeptide segment with a motif similar to the GxxxG dimerization motif of glycophorin A. Dimerization at the heptapeptide segment would be consistent with findings based on electrospray ionization MS data, chemical cross-linking studies, and CNBr cleavage data. 相似文献
156.
Visvaldas Kairys Michael K Gilson Burkhard Luy 《European journal of biochemistry》2004,271(11):2086-2092
The pulmonary surfactant prevents alveolar collapse and is required for normal pulmonary function. One of the important components of the surfactant besides phospholipids is surfactant-associated protein C (SP-C). SP-C shows complex oligomerization behavior and a transition to beta-amyloid-like fibril structures, which are not yet fully understood. Besides this nonspecific oligomerization, MS and chemical cross-linking data combined with CD spectra provide evidence of a specific, mainly alpha-helical, dimer at low to neutral pH. Furthermore, resistance to CNBr cleavage and dual NMR resonances of porcine and human recombinant SP-C with Met32 replaced by isoleucine point to a dimerization site located at the C-terminus of the hydrophobic alpha-helix of SP-C, where a strictly conserved heptapeptide sequence is found. Computational docking of two SP-C helices, described here, reveals a dimer with a helix-helix interface that strikingly resembles that of glycophorin A and is mediated by an AxxxG motif similar to the experimentally determined GxxxG pattern of glycophorin A. It is highly likely that mature SP-C adopts such a dimeric structure in the lamellar bilayer systems found in the surfactant. Dimerization has been shown in previous studies to have a role in sorting and trafficking of SP-C and may also be important to the surfactant function of this protein. 相似文献
157.
Eutrophication history of Lake Arendsee (Germany) 总被引:4,自引:0,他引:4
Burkhard W. Scharf 《Palaeogeography, Palaeoclimatology, Palaeoecology》1998,140(1-4):85-96
At the maximum depth of Lake Arendsee (49.5 m) only a thanatocenosis could be found in a freeze core with a length of 52 cm. According to varve-counting, the fauna changed between 1960 and 1972, characteristic of mesotrophic to eutrophic state. The cause for eutrophication could be correlated with the sewage loading of the town of Arendsee and the drainage of Lake Fauler See into Lake Arendsee. Lake Arendsee did not yet recover from the loading because of the long retention time of 114 years. The investigation of the living meio- and macrobenthos of Lake Arendsee shows that the lower profundal is not colonized. 相似文献
158.
Prongidi-Fix L Sugawara M Bertani P Raya J Leborgne C Kichler A Bechinger B 《Biochemistry》2007,46(40):11253-11262
The designed alpha-helical amphipathic peptide LAH4 assembles several properties, which makes it an interesting candidate as a gene-delivery vehicle. Besides being short and soluble in aqueous solutions, LAH4 presents cationic residues, which allow for efficient complexation of DNA. In addition, this peptide is poorly hemolytic at neutral pH, while it is able to destabilize biological membranes in acidic conditions. In this study, the structure of the peptide/DNA transfection complex was examined by circular dichroism and solid-state nuclear magnetic resonance spectroscopies and the thermodynamics of its formation and disassembly was monitored in a quantitative manner as a function of pH by isothermal titration calorimetry. Notably, the number of peptides within the complex considerably decreases upon acidification of the medium. This observation has direct and important consequences for the mechanism of action because the acidification of the endosome results in high local concentrations of free peptide in this organelle. Thus, these peptides become available to interact with the endosomal membranes and thereby responsible for the delivery of the transfection complex to the cytoplasm. When these data are taken together, they indicate a dual role of the peptide during the transfection process, namely, DNA complexation and membrane permeabilization. 相似文献
159.
Katrin Hayess Regine Kraft Jana Sachsinger Jürgen Janke Georg Beckmann Klaus Rohde Burkhard Jandrig Rainer Benndorf 《Journal of cellular biochemistry》1998,69(3):304-315
Recently, interest has focused on the human gene encoding the putative protein homologous to VAT-1, the major protein of the synaptic vesicles of the electric organ of the Pacific electric ray Torpedo californica, after it has been localized on chromosome locus 17q21 in a region encompassing the breast cancer gene BRCA1. Chromosomal instability in this region is implicated in inherited predisposition for breast and ovarian cancer. Here we describe isolation and biochemical characterization of a mammalian 48 kDa protein homologous to the VAT-1 protein of Torpedo californica. This VAT-1 homolog was isolated from a murine breast cancer cell line (Ehrlich ascites tumor) and identified by sequencing of cleavage peptides. The isolated VAT-1 homolog protein displays an ATPase activity and exists in two isoforms with isoelectric points of 5.7 and 5.8. cDNA was prepared from Ehrlich ascites tumor cells, and the murine VAT-1 homolog sequence was amplified by polymerase chain reaction and partially sequenced. The known part of the murine and the human translated sequences share 97% identity. By Northern blots, the size of the VAT-1 homolog mRNA in both murine and human (T47D) breast cancer cells was determined to be 2.8 kb. Based on the presented data, a modified gene structure of the human VAT-1 homolog with an extended exon 1 is proposed. VAT-1 and the mammalian VAT-1 homolog form a subgroup within the protein superfamily of medium-chain dehydrogenases/reductases. J. Cell. Biochem. 69:304–315, 1998. © 1998 Wiley-Liss, Inc. 相似文献
160.
Burkhard Bechinger 《Proteins》1997,27(4):481-492
The polypeptide corresponding to the signal sequence of the M13 coat protein and the five N-terminal residues of the mature protein was prepared by solid-phase peptide synthesis with a 15N isotopic label at the alanine-12 position. Multidimensional solution NMR spectroscopy and molecular modeling calculations indicate that this polypeptide assumes helical conformations between residues 5 and 20, in the presence of sodium dodecylsulfate micelles. This is in good agreement with circular dichroism spectroscopic measurement, which shows an α-helix content of approximately 42%. The α-helix comprises an uninterrupted hydrophobic stretch of ≤12 amino acids, which is generally believed to be too short for a stable transmembrane alignment in a biological bilayer. The monoexponential proton-deuterium exchange kinetics of this hydrophobic helical region is characterized by half-lives of 15–75 minutes (pH 4.2, 323 K). When the polypeptide is reconstituted into phospholipid bilayers, the broad anisotropy of the proton-decoupled 15N solid-state NMR spectroscopy indicates that the hydrophobic helix is immobilized close to the lipid bilayer surface at the time scale of 15N solid-state NMR spectroscopy (10−4 seconds). By contrast, short correlation times, immediate hydrogen-deuterium exchange as well as nuclear Overhauser effect crosspeak analysis suggest that the N and C termini of this polypeptide exhibit a mobile random coil structure. The implications of these structural findings for possible mechanisms of membrane insertion and translocation as well as for membrane protein structure prediction algorithms are discussed. © 1997 Wiley-Liss Inc. 相似文献