Analysis of glycopeptides as borate complexes by polyacrylamide gel electrophoresis

A new method using polyacrylamide gel electrophoresis in a Tris-borate buffer to analyze Pronase-derived glycopeptides is described. Examination of immunoglobulin, Sindbis virus, and ovalbumin-radiolabeled glycopeptides by this system demonstrates a pattern similar to that seen after Bio-Gel-P-6 chromatography and, in addition, exposes a heterogeneity in the immunoglobulin and Sindbis virus glycopeptides not apparent after gel filtration. The resolution of glycopeptides by gel electrophoresis depends on the inclusion of borate ions in the sample, the gel, and the electrophoresis buffer. The borate ions react with neutral sugars, converting them to charged complexes which migrate during electrophoresis. The number of borate ions bound to a glycopeptide is a function of the composition, sequence, and linkages of the carbohydrates. Gel electrophoresis of glycopeptides in a borate buffer has several advantages: (1) The method requires no new equipment or special skills beyond those necessary for conventional polyacrylamide gel electrophoresis; (2) when performed on a slab gel, up to 24 samples can be analyzed simultaneously; and (3) since detection is by radio-autography, small amounts of radiolabeled glycopeptides can be visualized by prolonging the exposure time. These characteristics are advantageous for studies of glycopeptides based on digestion products resulting from incubations with specific exo- and endo-glycosidases. Untreated glycopeptides have been compared on the same gel with glycopeptides sequentially treated with different glycosidases to gain structural information.     

Changes in Wheat Leaf Polysomal Messenger RNA Populations during the Early Stages of Rust Infection: EFFECTS OF CHLORAMPHENICOL AND LINCOMYCIN ON CELL-FREE TRANSLATION BY POLYSOMES FROM HEALTHY AND INFECTED LEAVES

Polysomes isolated from a susceptible variety of wheat leaves (cultivar W2691) and those inoculated with the wheat stem rust fungus (f. sp. tritici, race 126-ANZ-6, 7) were incubated in a cell-free protein-synthesizing system. Under these conditions, different size classes of polypeptides, ranging in molecular weight from 10,000 to 80,000, are radiolabeled. Using double-isotope labeling technique, we show that some discrete size classes of polypeptides are synthesized in significantly greater quantitites by polysomes from inoculated leaves compared to the corresponding size classes synthesized by polysomes from healthy leaves. These results confirm our previous observation that there are significant changes in the wheat leaf polysomal messenger RNA populations at 3 days after inoculation with the rust fungus.The effects of the organelle-specific inhibitors of protein synthesis, chloramphenicol and lincomycin, on in vitro polysomal messenger RNA translation were investigated. The polypeptides synthesized by polysomes from healthy and inoculated leaves in the presence of chloramphenicol were compared. The results show that, even in the presence of this antibiotic, the polysomes from inoculated leaves synthesize greater quantities of some size classes of polypeptides. These data indicate that changes in polysomal messenger RNA populations involve, at least in part, cytoplasmic messenger RNA.     

Inhibition of cyclopropane fatty acid synthase by sinefungin and A9145C

Sinefungin and A9145C, antifungal antibiotic analogs of S-adeno-sylmethionine isolated from $\text{Streptomyces}\text{,}\text{griseolus}$, have been found to be very effective $\text{in}\text{,}\text{vitro}$ inhibitors of cyclopropane fatty acid synthase from $\text{Lactobacillus}\text{,}\text{plantarum}$. Both compounds exhibit linear competitive inhibition with a K_i for Sinefungin of 220 nM and a K_i for A9145C of 11 nM.     

Steroid sulphatase levels in XX males,including observations on two affected cousins

Summary Quantitative assays of steroid sulphatase in XX males have shown that some individuals have two functional loci, and others only one. Two affected cousins, who cannot share the same X-chromosome, nevertheless have male levels of steroid sulphatase, suggesting functional abnormality of the X chromosome.The hypothesis is advanced that these and other unusual features of X-chromosome function in some XX males, could be explained if such cases were due to an autosomal mutation, exercising its effect by causing abnormal inactivation of a subterminal area of Xp which normally escapes the inactivation process.     

Variabilin,a 6a-hydroxypterocarpan from Dalbergia variabilis

Bark and wood of the creeper Dalbergia variabilis contain the previously described friedelin, O-acetyl-oleanolic acid, formononetin, 8-O-methylretusin, (+)-vestitol, (±)-mucronulatol, (+)- and (±)-medicarpin, besides (+)-variabilin [(6aR,11aR)-6a-hydroxy-3,9-dimethoxypterocarpan]. This structure was confirmed by the conversion of (+)-variabilin into di-O-methylcoumestrol.     

Susceptibility of house flies (Diptera: Muscidae) and five pupal parasitoids (Hymenoptera: Pteromalidae) to abamectin and seven commercial insecticides.

Assays of five commercial insecticides applied as residual sprays at label rates to plywood indicated the most toxic insecticide overall for pteromalid parasitoids of house flies, Musca domestica L., was Atroban (permethrin), followed by Ciodrin (crotoxyphos), Rabon (tetrachlorvinphos), Ectrin (fenvalerate), and Cygon (dimethoate). Insecticide-susceptible house flies were susceptible to all five insecticides (mortality, 62-100%). Flies that were recently colonized from populations on dairy farms in New York were susceptible only to Rabon. Urolepis rufipes (Ashmead) was the most susceptible parasitoid species overall to these insecticides, followed by Muscidifurax raptor Girault & Sanders, Nasonia vitripennis Walker, Pachycrepoideus vindemmiae (Rondani), and Spalangia cameroni Perkins. Compared with susceptible flies, newly colonized flies showed moderate resistance to avermectin B1a (abamectin). Abamectin was more toxic to all of the parasitoids except N. vitripennis and S. cameroni than to newly colonized house flies when exposed for 90 min to plywood boards treated with 0.001-0.1% abamectin. Space sprays with Vapona (dichlorvos) killed all of the parasitoids and susceptible flies and 64% of the newly colonized flies when insects were placed directly in the path of the spray; mortality was substantially lower among flies and parasitoids protected under 5 cm of wheat straw. Space sprays with Pyrenone (pyrethrins) killed greater than 86% of all insects exposed to the spray path except for the newly colonized flies (1% mortality); mortality of insects protected under straw was low (less than 12%) except for S. cameroni (76%). Because responses of the five parasitoids to the different insecticides varied considerably, general conclusions about parasitoid susceptibility to active ingredients, insecticide class, or method of application were not possible.     

Proteolytic degradation of heterologous proteins expressed inAspergillus niger

Summary Proteolytic degradation of heterologous proteins expressed in the filamentous fungusAspergillus niger reduces the yield of authentic target protein. The activities ofA. niger proteases are differentiated by their effects on two proteins expressed and secreted fromA. niger: hen egg-white lysozyme and porcine pancreatic phospholipase A₂.