C nuclear magnetic resonance studies of the binding of alkyl isocyanides to soybean leghemoglobin; comparison with animal myoglobins

¹³C NMR of labelled alkyl isocyanide ligands has been used with a view to probe the protein environment around the heme site of Soybean leghemoglobin, and comparatively, those of sperm whale myoglobin and monomeric Glycera hemoglobin. The terminal carbon of the isocyanide, which is known to be highly sensitive to change in hybridization of the nitrogen, could be expected to reflect the movement of the alkyl group through steric interactions. Three alkyl isocyanides (alkyl = methyl, ethyl & n-butyl) have therefore been used and the ¹³C° chemical shift values were measured for each ligand bound to the various proteins studied.In all cases, the ¹³C° resonances of the bound ligand were shifted considerably downfield with respect to those of the free unbound species, but the pattern of these displacements revealed more pronounced steric hindrance in the case of some proteins compared to others. The modifications of the chemical shift values on binding Δδ = δ_bound — δ_free) were least in the case of leghemoglobin; moreover, the Δδ values were insensitive to the length of the alkyl chain (methyl to n-butyl) when bound to leghemoglobin, in contrast to the other proteins examined. The results are interpreted as arising from a diminished steric hindrance to isocyanide binding with leghemoglobin, in conformity with the recently published X-ray structure which reports the existence of a large heme pocket on the distal side.     

Inhibition of glycolysis by furfural in Saccharomyces cerevisiae

Summary Furfural, a Maillard reaction product, was found to inhibit growth and alcohol production by Saccharomyces cerevisiae. Furfural concentrations above 1 mg ml^–1 significantly decreased CO₂ evolution by resuspended yeast cells. Important glycolytic enzymes such as hexokinase, phosphofructokinase, triosephosphate dehydrogenase, aldolase and alcohol dehydrogenase were assayed in presence of furfural. Dehydrogenases appeared to be the most sensitive enzymes and are probably responsible for the observed inhibition of alcohol production and growth.     

Biochemical studies on the origin of the ATPase of the avian myeloblastosis virus

The adenosine triphosphatase activity of the avian myeloblastosis virus obtained from the blood of the virus-infected chicken was compared with that of the host cell myeloblasts. The specific activity of the viral enzyme is unusually higher than that of the myeloblasts. A significant difference in inhibitor sensitivity was observed with quercetin. When the virus was grown in chicken embryonic fibroblasts in culture, the resulting virus showed very little adenosine triphosphatase activity, comparable to that of the fibroblasts and similar sensitivity to inhibitors. Antibody raised against the purified enzyme of avian myeloblastosis virus inhibits the enzyme activity of the myeloblasts while the activity of the fibroblasts enzyme as well as that of fibroblast-grown virus remains unaffected.     

alpha-adrenergic receptor in rat heart. Effects of thyroidectomy.

The effects of thyroid status on alpha-adrenergic receptors in the rat myocardium were investigated. The potent antagonist [3H]dihydroergokryptine was used to identify alpha-adrenergic receptors in rat heart particulate and sarcolemmal fractions. Administration of triiodothyronine to thyroidectomized rats decreased specific binding to alpha-adrenergic receptors in heart particulate and sarcolemmal fractions by 41% and 45%, respectively. Scatchard analysis revealed that the cardiac sarcolemmal fraction from thyroidectomized rats contained 29.3 fmol/mg of protein, as compared with 17.0 fmol/mg of protein found in the heart preparation of thyroidectomized rats treated with triiodothyronine. The equilibrium dissociation constants for the interaction of receptors with dihydroergokryptine were similar (about 1.5 nM) in the heart sarcolemmal fractions derived from these two groups of rats. The results of this study demonstrate that thyroid hormone can regulate the number of cardiac alpha-adrenergic receptors. In addition, there appears to be a reciprocal relationship between alpha-adrenergic and beta-adrenergic receptors in the rat myocardium.     

Complete nucleotide sequence of the leader rna synthesized in vitro by vesicular stomatitis virus

The complete nucleotide sequence of the leader RNA synthesized in vitro by the Indiana serotype of vesicular stomatitis virus is presented. The sequence was determined by the technique described by Donis-Keller, Maxam and Gilbert (1977) in combination with the standard two-dimensional fingerprint techniques described by Barrell (1971). The leader RNA contains 48 nucleotides variably terminating at the 3′ terminus with cytosine (68%) and adenosine at position 47 (32%). Since the leader RNA is complementary to the 3′ terminal portion of the viral genome RNA, the first 48 nucleotides from the 3′ end of the genome RNA can be deduced. The leader RNA contains repetitive and palindromic sequences with a polypurine sequence at its 3′ terminus. The possible role of some of the sequences is discussed.     

Hemagglutinating activity in extracts of mycelia from submerged mushroom cultures.

Extracts from mycelia of seven different mushrooms agglutinated erythrocytes of several species. More than one agglutinating factor was identified in the extracts of three different mycelia. Agglutination was partially inhibited nonspecifically by high concentrations of glucose, galactose, mannose, fucose, and rhamnose.     

Immobilization of yeast cells containing beta-galactosidase

Cultural conditions optimum for beta-galactosidase production by Saccharomyces anamensis are pH 4.5, temperature 26 +/- 2 degrees C, and 30 h of incubation period. Addition of lactose at 24 h fermentation greatly increase the level of enzyme. Optimum pHl, temperature, pH stability, and thermostability of yeast beta-galactosidase are negligibly affected by immobilization. The K(m) values of enzyme in the native and immobilized cells are 102mM and 148mM, respectively. Glucose noncompetitively inhibits the enzyme activity. Addition of substances such as dithioerythritol, glutathione, and bovine serum albumin to the native cell during assay procedure and immobilized cell prior to immobilization have stimulatory effects on enzyme activity. Metal ions like Ca(2+), Mg(2+) enhance the beta-galactosidase activity for both intact and bound cells. Immobilized cells retain 68.6% of the beta-galactosidase activity of intact cells and there is no significant loss of activity on storage at 4 degrees C for 28 days.     

Preparation and X-ray characterization of four new crystal forms of jacalin, a lectin from Artocarpus integrifolia

Four new crystal forms of the anti-T lectin from jackfruit (Artocarpus integrifolia) have been prepared and characterized. Three of them, two monoclinic (P21, a = 59.4 A, b = 83.3 A, c = 63.5 A, beta = 107.7 degrees; C2, a = 106.1 A, b = 53.9 A, c = 128.0 A, beta = 95.0 A) and one orthorhombic (C222(1), a = 98.1 A, b = 67.3 A, c = 95.1 A) were grown with 2-methylpentan-2,4-diol (MPD) as the precipitant while the fourth, an hexagonal form (P6(1)22, a = b = 129.6 A, c = 157.9 A), was obtained in the presence of methyl-alpha-D-galactopyranoside with polyethylene glycol 4000 as the precipitant. The reported relative molecular mass (Mr) of the lectin was found to be inconsistent with the solvent content of the crystals estimated using measured densities. The Mr was redetermined using size-exclusion chromatography in the presence of methyl-alpha-D-galactopyranoside and Ferguson-plot analysis of mobilities in polyacrylamide gel electrophoresis. The redetermined Mr (66,000) is consistent with the measured crystal densities. The orthorhombic and the hexagonal forms, which have one half molecule and one molecule, respectively, in the asymmetric unit, are suitable for high-resolution X-ray analysis.