Synthesis and application of acid labile anchor groups for the synthesis of peptide amides by Fmoc-solid-phase peptide synthesis

The preparation and application of a new linker for the synthesis of peptide amides using a modified Fmoc-method is described. The new anchor group was developed based on our experience with 4,4'-dimethoxybenzhydryl (Mbh)-protecting group for amides. Lability towards acid treatment was increased dramatically and results in an easy cleavage procedure for the preparation of peptide amides. The synthesis of N-9-fluorenylmethoxycarbonyl- ([5-carboxylatoethyl-2.4-dimethoxyphenyl)- 4'-methoxyphenyl]-methylamin is reported in detail. This linker was coupled to a commercially available aminomethyl polystyrene resin. Peptide synthesis proceeded smoothly using HOOBt esters of Fmoc-amino acids. Release of the peptide amide and final cleavage of the side chain protecting groups was accomplished by treatment with trifluoroacetic acid-dichloromethane mixtures in the presence of scavengers. The synthesis of peptide amides such as LHRH and C-terminal hexapeptide of secretin are given as examples.     

Cell dynamics and maturation within the olfactory epithelium proper of the mouse--a morphometric analysis

Principles of neurogenesis and neuronal maturation within theolfactory neuroepithelia are not fully understood. A morphometricanalysis, in which cell types were counted in BALB/c mice ofdifferent ages, was performed on the olfactory epithelium proper(OEP) in order to provide a clearer definition of these processes.Of particular interest were the density of mature receptor cells,the ratio between receptor cells and progenitor cells (lightbasal cells), and changes that occurred among basal cells withage. The results show that neurogenesis becomes slower withage, and suggest that some measure of mature receptor cell densitymay be important in controlling neurogenesis. The implicationsof these investigations to the development of topographicalprojections from the OEP to the olfactory bulb, and the establishmentof functional maps within the bulb are discussed.     

Synthesis of a monocharged peptide nucleic acid (PNA) analog and its recognition as substrate by DNA polymerases.

The preparation of a novel phosphoramidite monomer based on thyminyl acetic acid coupled to the secondary nitrogen of 2-(2-amino-ethylamino)ethanol is described. This monomer can be used to attach a deoxynucleotide to the carboxy terminus of a PNA oligomer by solid-phase synthesis. The resulting PNA primer is recognized as a substrate by various DNA polymerases.     

Synthesis of peptide amides by Fmoc-solid-phase peptide synthesis and acid labile anchor groups

The preparation and use of new anchor groups for the synthesis of peptide amides by solid-phase peptide synthesis employing the Fmoc-method is described. Based on the structure of the 4,4'-dimethoxybenzhydryl group (Mbh) handles were developed, which could be cleaved by mild acid treatment to give carboxamides. The syntheses and application of Fmoc-amino-acid-(4-carboxylatomethyloxyphenyl-4'-methoxyphenyl) methyl amide and Fmoc-(4-carboxylatopropyloxyphenyl-4'-methoxyphenyl) methyl amide are described in detail. These handles were coupled to resins and a stepwise elongation of peptide chains proceeded smoothly with N alpha-9-fluorenylmethoxycarbonyl (Fmoc) amino acid derivatives using a carbodiimide/HOBt mediated reaction. The final cleavage of side-chain protecting groups and the release of the C-terminal amide moiety was achieved by the treatment with trifluoroacetic acid, dichloromethane in the presence of scavengers. Various peptides, such as the Leu-enkephalin amide and Leu-Gly-Gly-Gly-Gln-Gly-Lys-Val-Leu-Gly-NH2, which is a good substrate for F XIII, were prepared in high yields and purities.     

Redox status of the eye lens

The aim of this work was to study the regional variation of some antioxidant systems in calf lens. Specific lens regions of nearly same age were obtained by a microsectioning technique, and the concentration of reduced and oxidized glutathione, protein sulfhydryl groups and iron were measured in each lens region. The concentration of reduced glutathione, the major redox buffer in lens, exponentially decreased from the cortical regions to the nucleus. In contrast, the concentration of protein sulfhydryl groups gradually increased from the cortex toward the nucleus. The protein-bound disulfides remained constant throughout the lens. Iron was concentrated in the outer cortical region. The results show that the most dynamic redox-active zone in the lens is the subcapsular cortical region where the oxidant flux meets a highly reducing environment containing a potent redox catalyst.     

Die Inkorporation von Leucin-H3 in Plexus chorioideus-Zellen in der Gewebekultur

Zusammenfassung Die Aufnahme tritiummarkierten Leucins von unterschiedlich reifen Plexus chorioideus-Zellen von Hühnerembryonen wird mit Hilfe der elektronenmikroskopischen Autoradiographie untersucht.Die frühen embryonalen Plexuszellen haben im Gegensatz zu den Entwicklungsstadien vor dem Schlüpfen noch kein endoplasmatisches Retikulum ausgebildet.In den Plexus chorioideus-Zellen 9 Tage alter Hühnerembryonen läßt sich eine ausgeprägte Aufnahme markierten Leucins in den Kern, den Golgiapparat und das freie Zytoplasma beobachten. Plexuszellen 17 Tage alter Hühnerembryonen zeigen gegenüber dem jüngeren Stadium eine verminderte Inkorporationsrate und eine nahezu selektive Lokalisation der Silbergrana über dem endoplasmatischen Retikulum im basalen Zellanteil.Die Verteilung des inkorporierten Materials im Kern und im Golgiapparat ist in den beiden Entwicklungsstadien vergleichbar. Die jüngeren Zellen inkorporieren eine größere Menge der markierten Aminosäure. Sie halten die markierte Substanz längere Zeit nach der Inkorporation zurück als ältere Elemente.Die Bedeutung des unterschiedlich schnellen Abbaus der markierten Verbindung in den untersuchten Entwicklungsstadien wird diskutiert.

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The uptake of leucine-H³ by choroid plexus cells in vitro as studied by electronmicroscopic autoradiography

Summary The uptake of tritium-labelled leucine by choroid plexus cells of the chick embryo during various stages of development was examined with electronmicroscopical autoradiography.In contrast to the developmental stage just prior to hatching, the early embryonic plexus cells do not possess an endoplasmic reticulum.The choroid plexus of the 9 days old chick embryo shows a marked uptake of labelled leucine by the nucleus, the Golgiapparatus and the free cytoplasm. Plexus cells of the 17 days old chick embryo show, unlike the earlier stages, a decreased incorporation and a near selective localization of the silver granules over the endoplasmic reticulum in the basal part of the cell.The distribution of the incorporated material in the nucleus and in the Golgiapparatus is comparable in both developmental stages examined. The younger cells incorporate a greater amount of labelled aminoacids.In comparison with the older cells younger cells retain the labelled aminoacids for a larger period after the incorporation.The significance of this difference for the metabolic rate of the labelled substances between the two developmental stages examined, is discussed.

Die Autoren danken Fräulein R. Dietrich, Frau D. Schulz und Frau M. C. Weinrichter für technische Assistenz.     

The vomeronasal epithelia of NMRI mouse

Summary The features of the apical and lateral surfaces of cells of the vomeronasal epithelium were studied in adult male mice by scanning electron microscopy. Supporting cells and receptor cells of the neuroepithelium are covered with microvilli. Microvilli of the sensory cells are longer and thinner than those of the supporting cells. Additionally, the former differ in local distribution, orientation, occurrence of branching and appearance of the cell coat. The receptor-free epithelium consists most likely of one cell type only, which shows different structural modifications including the presence, number and length of microvilli and cilia. In the transitional region, between the neuroepithelium and the receptor-free epithelium, immature receptor cells are present.This paper is dedicated to Prof. A. JinoSupported by grants from the Alexander von Humboldt-Stiftung and the Deutsche Forschungsgemeinschaft (Br. 358/5-1)     

Die Feinstruktur und Differenzierung des inneren Segmentes und des Paraboloids der Photorezeptoren in der Retina von Hühnerembryonen

Summary The development of the inner segment of the chick photoreceptors has been studied from the 6th to 21st day of incubation.The inner segment is essentially an elongation of the apical cytoplasm of the growing receptor, distally from the outer limiting membrane. An emigration of mitochondria follows, forming the ellipsoid.The paraboloid is a portion of the agranular endoplasmic reticulum and occupies a sharply localised non variable position within the receptor.Possible interrelatations between paraboloid, endoplasmic reticulum and Golgi apparatus are discussed. The presence of glycogen in the paraboloid seems to indicate that this specialised portion of e.r. may be either involved in glycolysis or a store for glycogen.

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Die Verfasser danken der Deutschen Forschungsgemeinschaft und der Volkswagenstiftung für ihre Unterstützung, Fräulein Ch. Kiele und Fräulein E. Möhring für die technische Assistenz, Frau M. Bothe für das Diagramm.