The application of a potential-sensitive cyanine dye to rat small intestinal brush border membrane vesicles

The sensitivity of the fluorescent dye, 3,3′-diethylthiadicarbocyanine (DiS-C₂(5)), was too low for the detection of membrane potential changes in rat small intestinal membrane vesicles. Only after adding LaCl₃ or after fractionation of the intestinal membranes by free-flow electrophoresis could the dye be used to monitor electrogenic Na⁺-dependent transport systems. It is concluded that the response of this potential-sensitive dye is influenced by the negative surface charge density of the vesicles.     

Intrinsic Bent DNA Sites in the Developmentally Amplified C3-22 Gene Promoter of Rhynchosciara americana (Diptera: Sciaridae)

The Rhynchosciara americana C3-22 gene is located in an amplified domain and is developmentally expressed. The aim of the present work was to identify intrinsically bent DNA sites in a segment containing the gene promoter and downstream sequence. The results indicated that this gene is flanked by intrinsically bent DNA sites. Three bent DNA sites (b^?3, b^?2, and b^?1) were localized in the promoter, and one was localized downstream of the gene (b⁺¹). These sites had helical parameters that confirmed the curved structure, as well as segments with left-handed superhelical writhe. In silico analysis of the promoters of four other insect genes, which encode secreted polypeptides, showed that they all had curved structures and similar helical parameters. Correlation with other results indicates that the detected intrinsically bent DNA sites that flank the C3-22 gene might be a consensus feature of the gene structure in the amplified domains.     

Suppression of Plant-parasitic Nematode Populations in Turfgrass by Application of Entomopathogenic Nematodes

We studied the influence of entomopathogenic nematodes , Steinernema carpocapsae and S. riobravis, on natural populations of plant - parasitic nematodes (PPNs) infesting turfgrass in Georgia and South Carolina . S. riobravis applied at 6 109 infective juveniles (IJs) / acre provided up to 95 - 100% control of the root - knot , Meloidogyne sp ., sting , Belonolaimus longicaudatus, and ring nematode , Criconemella sp ., in Georgia , but S. carpocapsae had no effect . S. riobravis was as effective as the chemical nematicide , Fenamiphos (Nemacur 10G) at 4 weeks after treatment and more effective at 8 weeks after treatment . In South Carolina , both S. riobravis and S. carpocapsae applied at 1 109 IJs / acre provided up to 86 - 100 % control of the root - knot , sting and ring nematodes . Application of 6 109 IJs / acre increased control by only 4 - 14 % over the 1 109 dosage . Possible causes of differences in efficacy of S. carpocapsae at the two sites are discussed . It is concluded that S. riobravis may provide effective , predictable and economical control of PPNs in turfgrass .     

Phospholipid dependence of the anion transport system of the human erythrocyte membrane. Studies on reconstituted band 3/lipid vesicles

Band 3 protein extracted from human erythrocyte membranes by Triton X-100 was recombined with the major classes of phospholipid occurring in the erythrocyte membrane. The resulting vesicle systems were characterized with respect to recoveries, phospholipid composition, protein content and vesicle size as well as capacity and activation energy of sulfate transport. Transport was classified into band-3-specific fluxes and unspecific permeability by inhibitors. Transport numbers (sulfate ions per band 3 per minute) served as a measure of functional recovery after reconstitution. The transport properties of band 3 proved to be insensitive to replacement of phosphatidylcholine by phosphatidylethanolamine, while sphingomyelin and phosphatidylserine gradually inactivated band-3-specific anion transport when present at mole fractions exceeding 30 mol%. The activation energy of transport remained unaltered in spite of the decrease in transport numbers. The results, which are discussed in terms of requirements of band 3 protein function with respect to the fluidity and surface charge of its lipid environment, provide a new piece of evidence that the transport function of band 3 protein depends on the properties of its lipid environment just as the catalytic properties of some other membrane enzymes. The well-established species differences in anion transport (Gruber, W. and Deuticke, B. (1973) J. Membrane Biol. 13, 19–36) may to some extent reflect this lipid dependence.     

Enzymatic O-glycosylation of kappa-caseinomacropeptide by ovine mammary Golgi membranes

The results of subcellular fractionation of sheep mammary gland membranes indicate that N-acetylgalactosaminyl polypeptide transferase and galactosyl-N-acetylgalactosaminyl transferase, which are involved in the assembly of disaccharide units of kappa-casein, are localized chiefly in Golgi membranes. The glycosyltransferase activities incorporating N-acetyl [1-14C] galactosamine and [U-14C] galactose from uridine diphosphate N-acetyl [1-14C] galactosamine and uridine diphosphate [U-14C] galactose, respectively, were measured after membrane solubilization with Triton X-100 either with unglycosylated caseinomacropeptide, or with this polypeptide containing the N-acetylgalactosamine side chain residues (desialylated and degalactosylated caseinomacropeptide). Radioactive N-acetylgalactosamine was incorporated in the unglycosylated acceptor peptide, and the glycosidic bonds in the product were alkali labile, suggesting that they were linked to the hydroxyamino acid residues. In addition radioactive N-acetylgalactosamine was released after alpha N-acetyl-D-galactosaminidase treatment of labelled caseinomacropeptide. [U-14C] galactose was incorporated in the desialylated and degalactosylated acceptor peptide. Reductive alkaline treatment of [U-14C] galactose peptide resulted in the release of a major product, the chromatographic properties of which in TLC were identical with authentic galactosyl (1 leads to 3) N-acetylgalactosaminitol. The structure of the labelled disacchariditol determined after periodate oxidation (two equivalents) by gas liquid chromatography-mass spectrometry revealed that the [U-14C] galactose was linked to position C-3 on the N-acetylgalactosaminyl-residue. The anomery of the galactose, as determined by a chemical method, indicates unambiguously a beta configuration.     

Chemical and physical methods in dating human skeletal remains

Chemical and physical methods for dating skeletal remains were examined. Benzidine reaction, ultra-violet fluorescence, specific gravity and supersonic conductivity were carried out on 71 dated skeletal findings distributed over the span of the last 3,500 years. Results given by benzidine reaction and ultra-violet fluorescence basically coincide, and positive readings were obtained up to about 200–350 years. Values measured in specific gravity and supersonic conductivity testing show a parallel trend, pointing out a clear difference between samples of the three first centuries and the ones belonging to more ancient periods examined.     

Identification of the 5-HT1A Receptor Binding Subunit in Rat Brain Membranes Using the Photoaffinity Probe [3H]8-Methoxy-2-[N-n-Propyl, N-3-(2-Nitro-4-Azidophenyl)Aminopropyl]Aminotetralin

The synthesis of a tritiated derivative of the 5-HT1A photoaffinity probe 8-methoxy-2-[N-n-propyl, N-3-(2-nitro-4-azidophenyl)aminopropyl]aminotetralin ([3H]8-methoxy-3'-NAP-amino-PAT) allowed the use of this probe for attempting the irreversible labeling of specific binding sites in rat brain membranes. Sodium dodecyl-sulfate-polyacrylamide gel electrophoresis of proteins solubilized from hippocampal microsomal membranes that had been incubated with 20 nM [3H]8-methoxy-3'-NAP-amino-PAT under UV light revealed a marked incorporation of 3H label into a 63-kilodalton protein termed PI. As expected of a possible correspondence between PI and 5-HT1A receptor binding sites, 3H labeling by the photoaffinity probe could be prevented by selective 5-HT1A ligands such as 8-hydroxy-2-(di-n-propylamino)tetralin, ipsapirone, buspirone, and gepirone and by N-ethylmaleimide, but not by the 5-HT2 antagonist ketanserin, noradrenaline- and dopamine-related drugs, monoamine oxidase inhibitors, and chlorimipramine. Furthermore, the regional and subcellular distributions of PI were identical to those of specific 5-HT1A binding sites. These results indicated that the binding subunit of the 5-HT1A receptor is a 63-kilodalton protein with a functionally important sulfhydryl group(s).