Biomedical subjects
Publications and source records attributed to K Burger.
Alpha-trifluoromethyl alpha-amino acids with unsaturated side-chains have been prepared from 5-fluoro-4-trifluoromethyloxazole and allyl, propargyl as well as terpene alcohols in a one-pot procedure.
5-Benzyloxy-4-trifluoromethyl-1,3-oxazoles, obtained from 5-fluoro-4-trifluoromethyloxazoles and benzyl alcohols, are capable for rearrangements. A 1,3 shift of a benzyl group is the key step of a new general route toward alpha-trifluoromethyl substituted aromatic and heteroaromatic amino acids, demonstrating that 5-fluoro-4-trifluoromethyl-1,3-oxazole is a synthetic Tfm-Gly equivalent. On reaction with benzpinacol partially fluorinated oxazoles are transformed into bis(trifluoromethyl) substituted 2,5-diamino adipic acid and N-benzoyl-2-benzhydryl-3,3,3-trifluoroalanine.
Amino acid sequences and linear or head-to-tail cyclopeptides can be represented conveniently in one-line text formulae using the three-letter symbols. However, other - but nonetheless important - topologies of peptides are 'side chain-to-head (or tail)', 'backbone-to-backbone', 'side chain-to-side chain' cyclopeptides, 'side chain-to-side chain' connected peptide strands, and branched peptides (like peptide dendrimers). In general, such structures cannot be described using the three-letter symbols in one-line text: a chemical structure editor is required for symbolic representations according to the IUPAC-IUBMB recommendations. The aim of this contribution is to offer an unambiguous and general nomenclature system that enables researchers to represent all cyclic and branched homo- and hetero-detic peptides in a coherent manner in one-line text - as long as their as constituents can be represented in (three)-letter codes. The application of this new nomenclature would overcome the existing difficulties and provide a way to express complex situations in the shortest way in order to highlight more clearly the salient points in a given scientific communication.
Hexafluoroacetone was applied as a bidentate protecting and activating agent for the syntheses of RGD-peptide mimetics starting from iminodiacetic acid in solution and on solid phase.
Mercury sphygmomanometers are being removed from clinical practice in the United States due to environmental concerns about mercury toxicity. Accurate blood pressure measurement is central to high-quality hypertension management. In this study of 106 patients, the BpTRU(TM) device was compared to nurse blood pressure measurements that complied with all the JNC VII/American Heart Association guidelines in evaluation of a random casual blood pressure. The intermethod difference in systolic blood pressure was +1.8+/-5.1 mmHg, and for diastolic blood pressure it was 4.8+/-5.1 mmHg (both P<0.001). For the primary study end point of clinical decision-making, there was 92% (97/106) agreement between the hypertension nurse specialist and the BpTRU (kappa 0.8280, 95% confidence interval, 0.721-0.9350). The oscillometric blood pressure measurement with the BpTRU is recommended as a replacement for poorly performed auscultatory blood pressure measurement in clinical practice.
Solvation of a tetrapeptide, NAc-Ser-Phe-Val-Gly-OMe (1), in water and in water/alcohol mixtures with 2,2,2-trifluoroethanol (TFE)/water or ethanol (ETH)/water has been studied by diffusion NMR and intermolecular NOE measurements. The experimental results were compared with those obtained from detailed Molecular Dynamics (MD) calculations. Independently, all three methods revealed preferential solvation on the surface of the peptide by TFE in the water/TFE mixtures, but not by ETH in the water/ETH mixtures. The MD calculations show that the TFE concentration coating the peptide is higher than that in the bulk, while for ethanol, the concentration is nearly equal to that in the bulk. Calculated site-specific preferential solvation data between TFE, ETH, and water with the different peptide groups have been compared with the NMR data and shown to be in general agreement with the experimental facts.
Bombesin is a tetradecapeptide that possesses a random coil structure in pure water. In the presence of 30 % (v/v) 2,2,2-trifluoroethanol (TFE), it adopts a partial helical conformation involving the C-terminal amino acids 6-14. This conformational change, known as the TFE effect, is studied here in terms of the solvation state of the peptide at different TFE concentrations by means of intermolecular homo- and heteronuclear NOE measurements. When an aqueous solution of bombesin is titrated with TFE, a continual decrease in the water/peptide interactions and a concomitant increase in the TFE/peptide interactions is observed, and at 30 % (v/v) TFE no homonuclear NOEs between water and the peptide can be detected. The conformational transition of the bombesin molecule is thus accompanied by a complete surface covering with TFE. A parallel molecular dynamics (MD) study of the peptide in aqueous solution with the single-point charge (SPC) water model and in a 30 % (v/v) TFE/water mixture with a recently developed TFE model has also been performed. The 10 ns simulations were in agreement with the experimental data. The calculations indicate stabilisation of the alpha-helix in the H(2)O/TFE mixture, in contrast to the situation in pure water, and clustering of the TFE molecules around the peptide.
The coordination of zinc(II) ion to hyaluronate (Hya), a natural copolymer, in aqueous solution at pH 6 was investigated by potentiometric and circular dichroism (CD) spectroscopic methods, and by monitoring the changes in macroscopic properties by high-precision measurements. The zinc(II)-selective electrode, and CD measurements proved the binding of zinc(II) by Hya. A number of Hya fragments (Mr approximately 3.3 x 10(3)-1.4 x 10(6)) were studied to estimate the contributions of the polyelectrolyte effect, the solvation and host-guest interactions to the extra stabilization of the macromolecular zinc(II) complexes as compared with the monomeric unit. The zinc(II) ion activity increase reflected a stability decrease for the fragments with Mr < 4 x 10(4). This molecular weight differs from that where cleavage of the Hya skeleton starts (approximately 5 x 10(5), according to the size-exclusion gel, and anion-exchange chromatographic behavior of the Hya fragments) and from that where the polyelectrolyte effect stops (approximately 6 x 10(3)). The excess volumes and Bingham shear yield values of the solutions revealed the transformation of the coherent random coil structure stabilized by intermolecular association in the NaHya to an intramolecular association producing the globular structure of the ZnHya molecule, with a smaller but more strongly bound solvate water sheet.
Recent data revealed that DEK associates with splicing complexes through interactions mediated by serine/arginine-repeat proteins. However, the DEK protein has also been shown to change the topology of DNA in chromatin in vitro. This could indicate that the DEK protein resides on cellular chromatin. To investigate the in vivo localization of DEK, we performed cell fractionation studies, immunolabeling, and micrococcal nuclease digestion analysis. Most of the DEK protein was found to be released by DNase treatment of nuclei, and only a small amount by treatment with RNase. Furthermore, micrococcal nuclease digestion of nuclei followed by glycerol gradient sedimentation revealed that DEK co-sedimentates with oligonucleosomes, clearly demonstrating that DEK is associated with chromatin in vivo. Additional chromatin fractionation studies, based on the different accessibilities to micrococcal nuclease, showed that DEK is associated both with extended, genetically active and more densely organized, inactive chromatin. We found no significant change in the amount and localization of DEK in cells that synchronously traversed the cell cycle. In summary these data demonstrate that the major portion of DEK is associated with chromatin in vivo and suggest that it might play a role in chromatin architecture.
The first synthesis of a series of totally orthogonal protected racemic alpha-(trifluoromethyl)- and alpha-(difluoromethyl)arginines is described. The key steps of the synthesis are the mild guanidinylation procedure and the selective hydrogenation of a CC triple bond in the presence of a Cbz-group.
Cholesterol influences many of the biophysical properties of membranes and is nonrandomly distributed between cellular organelles, subdomains of membranes, and leaflets of the membrane bilayer. In combination with the high dynamics of cholesterol distribution, this offers many possibilities for regulation of membrane-embedded receptors. Depending on the receptor, cholesterol can have a strong influence on the affinity state, on the binding capacity, and on signal transduction. Most important, cholesterol may stabilize receptors in defined conformations related to their biological functions. This may occur by direct molecular interaction between cholesterol and receptors. In this review, we discuss the functional dependence of the nicotinic acetylcholine receptor as well as different G protein-coupled receptors on the presence of cholesterol.
Cholesterol affects the ligand binding function of the oxytocin receptor in a highly specific manner. While the structurally-related cholecystokinin receptor shows a strong correlation between the membrane fluidity and its binding function, the oxytocin receptor behaves differently. A stringent and unique requirement of the affinity state of the oxytocin receptor for structural features of the sterol molecule has been found. The molecular requirements differ both from those postulated for sterol-phospholipid interactions and from those known to be necessary for the activity of other proteins. Employing a new detergent-free subcellular fractionation protocol, a two-fold enrichment of the oxytocin receptors (10-15% of total receptors) has been detected in the cholesterol-rich, caveolin-containing membrane domains of the plasma membrane. While most of the properties of the oxytocin receptors were indistinguishable in cholesterol-poor versus cholesterol-rich membrane compartments, high-affinity oxytocin receptors localised in caveolin-enriched low-density membranes showed about a 3-fold higher stability against thermal denaturation at 37 degrees C compared with the oxytocin receptors localised in high-density membranes. Moreover, addition of cholesterol to the cholesterol-poor high-density membranes fully protected the oxytocin receptors against thermal denaturation and partially rescued high-affinity oxytocin binding. Although the membrane fluidity of the caveolin-enriched domains was lower than that in the high-density membranes, there was no correlation between the stability of oxytocin receptors and the fluidity level of the membrane domains. Finally, in a molecular modelling approach a putative cholesterol binding motif on the extracellular surface of the oxytocin receptor was found.
OBJECT: The outcome for children with recurrent malignant brain tumors is poor. The majority of patients die of progressive disease within months of relapse, and other therapeutic options are needed. The goal of this Phase I study was to evaluate the safety of in vivo suicide gene therapy in 12 children with recurrent, malignant, supratentorial brain tumors. METHODS: After optimal repeated tumor resection, multiple injections of murine vector-producing cells shedding murine replication-defective retroviral vectors coding the herpes simplex virus thymidine kinase type 1 (HSV-Tk1) gene were made into the rim of the resection cavity. Fourteen days after the vector-producing cells were injected, ganciclovir was administered for 14 days. The retroviral vector that was used only integrated and expressed HSV-Tk1 in proliferating cells, which are killed after a series of metabolic events lead to cell death. The median age of the patients was 11 years (range 2-15 years). Treated brain tumors included seven malignant gliomas, two ependyminomas, and three primitive neuroectodermal tumors. The patients were treated with one of three escalating dose concentrations of vector-producer cells. Four transient central nervous system adverse effects were considered possibly related to the vector-producing cells. In no child did permanent neurological worsening or ventricular irritation develop, and tests for replication-competent retroviruses yielded negative findings. CONCLUSIONS: This Phase I study demonstrates that in vivo gene therapy in which a replication-defective retroviral vector in murine vector-producing cells is delivered by brain injections can be performed with satisfactory safety in a select group of children with localized supratentorial brain tumors.
Progesterone at concentrations between 10 microM and 200 microM affected the calcium signaling evoked by ligand stimulation of G protein-coupled receptors expressed in several cell lines. At 160 microM progesterone the signaling of all receptors was completely abolished. The effect of progesterone was fast, reversible and was not prevented by cycloheximide indicating its non-genomic nature. Overall, the action of progesterone was more cell type-specific than receptor-specific. Our results are in contrast to a recent report [Grazzini, E., Guillon, G., Mouillac, B. and Zingg, H.H. (1998) Nature 392, 509-512] in which a direct high-affinity interaction between the oxytocin receptor and progesterone was suggested.
Anomalous scattering of X-rays at a synchrotron source can be used for the ab initio structure determination of unknown crystal structures using only powder diffraction data. For noncentrosymmetric crystals, the phases of structure factors can only be determined with a remaining ambiguity, when one chemical element is used as resonant scatterer. A corresponding additional constraint function has been built into an enhanced version of the program MEED, so that now all types of information gained from an anomalous-scattering powder diffraction experiment can be used in a maximum-entropy calculation of the electron-density distribution: phased reflections, unphased reflections, intensities of groups of overlapping reflections, and now also reflections with a remaining ambiguity in the phase. This is important for practical use, since a lot of information is already lost in the powder diagram compared with single-crystal datasets and it is essential to use all remaining information. The new constraint is demonstrated with the structure of Cu(5)Zn(8).
The translocation of spin-labeled analogues of phosphatidylcholine (4-doxylpentanoyl-PC, SL-PC), phosphatidylethanolamine (SL-PE), phosphatidylserine (SL-PS), and sphingomyelin (SL-SM) from the outer to the inner leaflet of the plasma membrane bilayer was investigated in dog kidney MDCK II and human colon Caco-2 cells. Disappearance from the outer leaflet was assayed using back-exchange to serum albumin. Experiments with cells in suspension as well as with polarized cells on filters were performed at reduced temperatures (10 and 20 degreesC) to suppress endocytosis and hydrolysis of spin-labeled lipids. For both epithelial cell lines, a fast ATP-dependent inward movement of the aminophospholipids SL-PS and SL-PE was found, while SL-SM was only slowly internalized without any effect of ATP depletion. The kinetics of redistribution of SL-PC were clearly different between the two cell lines. In MDCK II cells, SL-PC was rapidly internalized in an ATP-dependent and N-ethylmaleimide-sensitive manner and at a rate similar to that of the aminophospholipids. In contrast, in Caco-2 cells the inward movement of SL-PC was much slower than that of the aminophospholipids, did not depend on ATP, and was not N-ethylmaleimide-sensitive. Inhibitor studies indicated that the outward-translocating multidrug resistance P-glycoprotein present in these cells did not affect the kinetics of inward translocation. Internalization was always similar on the apical and basolateral cell surface, suggesting the presence of the same phospholipid translocator(s) on both surface domains of epithelial cells. We propose that Caco-2 cells contain the well-known aminophospholipid translocase, while MDCK II cells contain either two translocases, namely, the aminophospholipid translocase and a phosphatidylcholine-specific translocase, or one translocase of a new type, translocating aminophospholipids as well as phosphatidylcholine.
A new method for the preparation of N-methylamino acids and some of their derivatives starting from hexafluoroacetone protected amino acids is described. The new concept results in saving of steps compared to conventional protection/activation techniques. Protection and deprotection proceed without recemization.
To study the effect of the conformation of sugar hydroxy groups on metal complexation processes, complex formation of eight saccharides (D-fructose, L-sorbose, L-arabinose, D-arabinose, D-glucose, D-sorbitol, 2-deoxy-D-glucose and D-saccharose) with dimethyltin(IV)2+ cations was investigated in aqueous solution by potentiometric equilibrium measurements, 13C NMR, polarimetric and Mössbauer spectroscopic methods. The experimental results proved that deprotonation of D-fructose and L-sorbose is caused by the coordination of dimethyltin(IV)2+ in the unusual low pH interval 4-6 in contrast to the other saccharides deprotonated in analogous way at pH > 8. Increasing the pH of the solution resulted in the formation of further complexes. Stability and composition of the species was determined by potentiometric studies. 13C NMR measurements led to the assignment of the sugar OH groups participating in the processes. Mössbauer investigations in the quick-frozen solutions permitted the determination of the stereochemistry of tin(IV) in the complexes.