Solubilization of the active vasoactive intestinal peptide receptor from human colonic adenocarcinoma cells

The non-ionic detergent n-octyl-beta-D-glucopyranoside was used to solubilize the VIP (vasoactive intestinal peptide) receptor from human colonic adenocarcinoma cell line HT29-D4. The binding of monoiodinated 125I-VIP to the solubilized receptor was specific, time-dependent, and reversible. Scatchard analysis of data obtained from competitive displacement of monoiodinated 125I-VIP by native VIP suggested the presence of two classes of VIP binding sites with Kd values of 0.32 and 46.7 nM. The binding capacities of these two classes were 1.7 x 10(10) and 30.2 x 10(10) sites/mg of proteins, respectively. The solubilized receptor retained the specificity of the human VIP receptor towards the peptides of the VIP/secretin/glucagon family. The order of potency in inhibiting monoiodinated 125I-VIP binding was VIP (IC50 = 1.0 x 10(-9) M) much greater than peptide histidine methionine amide (IC50 = 10(-7) M) greater than growth hormone-releasing factor (IC50 = 3 x 10(-7) M) greater than secretin (IC50 greater than 10(-6) M); glucagon had no effect on VIP binding. The reducing agent dithiothreitol inhibited in a dose-dependent manner the binding of 125I-VIP. Covalent cross-linking experiments between the solubilized receptor and 125I-VIP showed that after sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography two major and one minor polypeptides of Mr 67,000, 72,000, and 83,000 were specifically labeled. When analyzed by gel filtration, the n-octyl-beta-D-glucopyranoside-solubilized 125I-VIP-receptor complex was resolved into two major peaks with molecular mass in the range of 60-70 and 270-300 kDa. Thus, the soluble form of the VIP receptor was probably a multimeric complex in which disulfide bonds may play an important role to hold the receptor in an active configuration.     

Saprophytic production of Clavine alkaloids and activity of Hydroxymethylglutaryl-CoA reductase

In submerged Claviceps cultures the activity of hydroxymethylglutaryl-CoA reductase preceded the increase of alkaloid production and of sterol content. During the first alkaloid phase, cell mevalonate was involved in the biosynthesis of both alkaloids and steroids. In the second production phase, it was predominantly used for alkaloid synthesis. Hydroxymethylglutaryl-CoA reductase appears to be a suitable target for physiological manipulation to increase clavine alkaloid yields.     

Mutations affecting the enzymes involved in the utilization of 4-aminobutyric acid as nitrogen source by the yeast Saccharomyces cerevisiae

We present genetic evidence for the enzymes 4-aminobutyrate: 2-oxoglutarate aminotransferase (EC 2.6.1.19) and succinate-semialdehyde dehydrogenase [NAD(P)+] (EC 1.2.1.16) constituting the functional pathway for the utilization of 4-aminobutyric acid as a nitrogen source by Saccharomyces cerevisiae. We show that the pathway is induced by 4-aminobutyric acid and that the presence of the pathway enzymes probably requires the integrity of a positive control element.     

Effects of histamine on bronchial artery blood flow and bronchomotor tone

The effects of aerosolized 5% histamine (10 breaths) on bronchial artery blood flow (Qbr), airflow resistance (RL), and pulmonary and systemic hemodynamics were studied in mechanically ventilated sheep anesthetized with pentobarbital sodium. Histamine increased mean Qbr and RL to 252 +/- 45 and 337 +/- 53% of base line, respectively. This effect was significantly different from base line for 30 min after challenge. The histamine-induced increase in RL was blocked by pretreatment with the histamine H1 receptor antagonist, chlorpheniramine, whereas the histamine-induced elevation in Qbr was prevented by the H2 antagonist, metiamide. Both responses were blocked only when both antagonists were present. Changes in Qbr were not directly associated with alterations in systemic and pulmonary hemodynamics or arterial blood gas composition. In vitro histamine caused a dose-dependent contraction of ovine bronchial artery strips that was prevented by H1 antagonist. The H2 agonist, impromidine, caused relaxation of precontracted arterial strips and was more potent and efficacious than histamine, whereas H1 agonists failed to elicit a relaxant response. Thus these findings indicate that histamine aerosol induces a vasodilation in the bronchial vascular bed; histamine has a direct effect on Qbr that is independent of alterations in RL, systemic and pulmonary hemodynamics, or arterial blood gas composition; and, histamine-induced bronchoconstriction is mediated predominantly by H1-receptors, whereas increased Qbr is controlled predominantly by H2-receptors, probably located in resistance vessels. This local effect of histamine on Qbr may have important implications in the pathophysiology of bronchial asthma and pulmonary edema.     

Growth of Streptomyces bambergiensis and glucose isomerase biosynthesis

Summary Growth and glucose isomerase biosynthesis in Streptomyces bambergiensis ATCC 13879 have been studied under different conditions. Some data concerning correlation between cultivation conditions and elemental analysis of the cells are also presented.     

Die submontanen krautreichen Buchenwälder auf Silikatböden der westlichen Tschechoslowakei

Die submontanen krautreichen Buchenwälder auf Silikatböden in der westlichen Tschechoslowakei (Tschechische Sozialistische Republik) zerfallen in vier Assoziationen: 1.Tilio cordatae-Fagetum Mráz 1960 em.Moravec 1977, 2.Tilio platyphylli-Fagetum Klika 1939, 3.Melico-Fagetum Seibert 1954 und 4.Carici pilosae-Fagetum Oberdorfer 1957. DasTilio cordatae-Fagetum ist an niedrigere Bergländer Mittel-, Süd- und Westböhmens gebunden. DasTilio platyphylli-Fagetum ist eine lokal bedingte Assoziation des Westteils des Gebirges ?eské st?edoho?í. DasMelico-Fagetum kommt in Nord- bis Ostböhmen und Nord- und Mittelmähren vor. DasCarici pilosae-Fagetum ist auf die Karpaten beschränkt (ausnahmsweise greift es auf den Ostrand des Böhmischen Hochlandes über). Die Buchenwälder des Hügellandes bei Jevany (Mittelböhmen) und des Gebirges ?elezné hory gehören, trotz dem Vorkommen in submontanen Lagen, demDentario enneaphylli-Fagetum Oberdorfer ex W. etA. Matuszkiewicz 1960 an. Die Zusammensetzung der Assoziationen ist durch pflanzensoziologische Tabellen belegt und ihre Verbreitung auf einer Punktkarte erfasst.     

Identification, characterization, and molecular cloning of a novel transporter-like protein localized to the central nervous system.

During the course of large scale purification of the D1 dopamine receptor from rat brain, a protein of approximately 87,000 daltons (p87) was observed to copurify with the D1 receptor through four chromatographic steps. To characterize the nature of this protein, bovine and rat cDNA clones were isolated and sequenced. The bovine and rat clones were highly conserved (98.5% identity). Each clone possessed an open reading frame of 2226 base pairs encoding a protein of 742 amino acids (calculated MW of 82,500), containing three stretches of peptide sequence obtained from p87 sequence analysis. Comparison of the deduced peptide sequence of this protein with those found in available databanks revealed that it was a novel protein related to the family of nutrient transport proteins from eukaryotes and bacteria, including, the mammalian facilitated glucose transporters, the yeast transporters for maltose, lactose, and glucose, and the proton-driven bacterial transporters for arabinose, xylose, and citrate. In addition p87 also shares with these transporters a similar hydropathicity profile that suggests the presence of 12 transmembrane segments. The mRNA for p87 appears to be localized primarily, if not exclusively, to the central nervous system. Northern blot analysis reveals a message of approximately 4.8 kb in cortex, hippocampus, brain stem, and cerebellum, but no detectable signal in peripheral tissues such as spleen, liver, kidney, lung, heart, or skeletal muscle. Evidence form Western blot analysis and immunohistochemistry suggests that this protein may be expressed in intracellular organelles or the membrane of synaptosomes rather than plasma membrane. Based on its structure and properties, p87 appears to define a new class of transporter-like proteins.