Nitrite reduction in Bacteroids of Rhizobium “hedysari” strain HCNT 1

Ex planta, bacteroids of the sulla-symbiont Rhizobium hedysari strain HCNT 1 terminated reduction of nitrite at nitrous oxide irrespective of the presence or absence of acetylene. Nitrate was not reduced during the experimental period, but slight nitrate reductase activity occurred if incubation with nitrate was prolonged (up to 15 h). As was observed in free-living cells, exposure of the bacteroids to the metal chelator, diethyldithiocarbamate, prevented reduction of nitrite, indicating the presence of a copper-containing nitrite reductase. Pulses of 10–75 M nitrite transiently impeded O₂ uptake in bacteroids, which resumed consumption of O₂ when the nitrite had been reduced. Exposure to >1.0 mM nitrite for 24h greatly inhibited nitrogenase activity (assayed as acetylene reduction activity) of bacteroids in planta. Exposure to the same concentrations of nitrite after 1h of incubation in the presence of acetylene almost completely stopped ongoing ethylene production in bacteroids of strain HCNT 1 extracted from nodules. Free cells of the non-nitrite-reducing R. hedysari strain CC 1335 were lacking in nitrogenase (acetylene-reduction) activity, whereas identically cultured (low-oxygen) strain HCNT 1 cells reduced both nitrite and acetylene.Abbreviations PMS phenazine methosulfate - DDC diethyldithiocarbamate     

A re-examination of the Na+-independent binding of [3H]β-alanine to rat brain stem-spinal cord

The Na⁺-independent binding of [³H]-alanine to rat brain stem plus spinal cord was reinvestigated, in order to study in more detail the characteristics of previously described -alanine binding processes. Binding was absent when amino acid-free postnuclear supernatants or crude synaptic membranes were used. Experiments performed with several other Na⁺-free preparations showed a sole binding component, irrespective of the preparation used. Biochemical characterization of this Na⁺-independent binding, using frozen/thawed/washed synaptosomal-mitochodrial fractions, showed that binding reached a plateau between 7 min and 13 min, increasing thereafter. Binding was linear with fraction protein over a range of 200–415 g/ml incubation medium. Binding was completely inhibited by glycine, alanine, -aminobutyric acid, -aminoisobutyric acid, hypotaurine and strychnine, and to a lesser extent by 2,2-dimethyl--alanine, brucine and gelsemine. It was insensitive to taurine, -aminobutyric acid (GABA), 2-guanidinoethanesulfonic acid (GES), carnosine, and bicuculline methiodide. Binding was reversible, saturable (K _D 20 M), and heat sensitive.     

A role for haemoglobin in all plant roots?

Abstract. We have found haemoglobin in plant roots whereas previously it has been recorded only in nitrogen fixing nodules of plants. Haemoglobin occurs not only in the roots of those plants that are capable of nodulation but also in the roots of species that are not known to nodulate. We suggest that a haemoglobin gene may be a component of the genome of all plants. The gene structure and sequence in two unrelated families of plants suggests that the plant haemoglobins have had a single origin and that this origin relates to the haemoglobin gene of the animal kingdom. At present we cannot completely rule out the possibility of a horizontal transfer of the gene from the animal kingdom to a progenitor of the dicotyledonous angiosperms but we favour a single origin of the gene from a progenitor organism to both the plant and animal kingdoms. We speculate about the possible functions of haemoglobin in plant roots and put the case that it is unlikely to have a function in facilitating oxygen diffusion. We suggest that haemoglobin may act as a signal molecule indicating oxygen deficit and the consequent need to shift plant metabolism from an oxidative to a fermentative pathway of energy generation.     

Proteolytic specificity of chicken cathepsin L on bovineβ-casein

The proteolytic specificity of chicken cathepsin L was studied using bovine -casein as substrate. The peptide mixtures obtained after various times of hydrolysis were separated by RP-HPLC and ten peptides were identified. Chicken cathepsin L accepts proline residues in all positions except P ₁ . Looking at the amino acid residues on the amino side of the scissile bond we found three times the Tyr-Pro pair at P ₁ –P ₂ positions and that the S ₁ subsite can interact with modified amino acids such as phosphoserine.Abbreviations RP-HPLC reverse phase high performance liquid chromatography - NMec N-methyl coumarylamide - TEA triethylamine - TFA trifluoroacetic acid     

Immobilization of enzymes on alumina by means of pyridoxal 5′-phosphate

A number of proteases have been immobilized on alumina in a two-step procedure: the first step converted them into semisynthetic phosphoproteins which, in the second step, spontaneously bonded to alumina through their phosphate function. The immobilized enzymes thus obtained showed the physical properties typical of the inorganic carrier and a high activity on low molecular weight substrates.     

A model for gramicidin A′-phospholipid interactions in bilayers

A model is proposed for the effect of gramicidin A on the order and structure of phospholipid dispersions. According to this model, the addition of gramicidin A influences the surrounding lipids via two independent mechanisms. The first arises from a drop in surface pressure for those lipids substantially bounded by gramicidin A. The second mechanism arises from the increase in the phospholipid headgroup spacing due to the small polar region of the polypeptide. The model provides an explanation for the currently available NMR, X-ray diffraction and Langmuir monolayer results. The model also suggests mechanisms for the ability of gramicidin A to trigger a transition of the lipid from the lamellar to hexagonal II phase, the dependence of this transition on the lipid chain length and the formation of a lamellar phase with lysophosphatidylcholine.Abbreviations NMR nuclear magnetic resonance - DMPC dimyristoylphosphatidylcholine - S molecular order parameter - CSA chemical shift anisotropy - DPPC dipalmitoylphosphati-dylcholine - LPC lysophosphatidylcholine     

Correlation of physical maps and some genetic functions in the genomes of the kappa-theta phage family of Bacillus licheniformis

Summary Natural recombinant genomes between several, phenotypically distinct forms of phages and were isolated and characterized by DNA restriction fragment mapping and electron microscopic heteroduplex analysis. The phenotypes of the recombinants were correlated with the physical maps of the genomes, and several genetic functions were therfore defined and mapped. All genes necessary for the assembly of infectious virus particles map in a contiguous tract of DNA comprising about 20 kb, or nearly one third of the genome length. No DNA homology occurs within these domains of the two genomes, so that homologous recombination does not take place here and phenotypic mixing of the phages is eo ipso excluded. Other regions of heterology contain regulatory genes responsible for the lytic or temperate character of the phages, and for exclusion of phage by .     

Voltage dependence of Na/K pump current inXenopus oocytes

Summary Stage V and VI (Dumont, J.N., 1972.J. Morphol. 136:153–180) oocytes ofXenopus laevis were treated with collagenase to remove follicular cells and were placed in K-free solution for 2 to 4 days to elevate internal [Na]. Na/K pump activity was studied by restoring the eggs to normal 3mm K Barth's solution and measuring membrane current-voltage (I–V) relationships before and after the addition of 10 m dihydroouabain (DHO) using a two-microelectrode voltage clamp. Two pulse protocols were used to measure membraneI–V relationships, both allowing membrane currents to be determined twice at each of a series of membrane potentials: (i) a down-up-down sequence of 5 mV, 1-sec stair steps and (ii) a similar sequence of 1-sec voltage pulses but with consecutive pulses separated by 4-sec recovery periods at the holding potential (–40 mV). The resulting membraneI–V relationships determined both before and during exposure to DHO showed significant hysteresis between the first and second current measurements at each voltage. DHO difference curves also usually showed hysteresis indicating that DHO caused a change in a component of current that varied with time. Since, by definition, the steady-state Na/K pumpI–V relationship must be free of hysteresis, the presence of hysteresis in DHO differenceI–V curves can be used as a criterion for excluding such data from consideration as a valid measure of the Na/K pumpI–V relationship. DHO differenceI–V relationships that did not show hysteresis were sigmoid functions of membrane potential when measured in normal (90mm) external Na solution. The Na/K pump current magnitude saturated near 0 mV at a value of 1.0–1.5 A cm^–2, without evidence of negative slope conductance for potentials up to +55 mV. The Na/K pump current magnitude in Na-free external solution was approximately voltage independent. Since these forward-going Na/K pumpI–V relationships do not show a region of negative slope over the voltage range –110 to +55 mV, it is not necessary to postulate the existence of more than one voltage-dependent step in the reaction cycle of the forward-going Na/K pump.     

Expression sites and developmental regulation of genes encoding (1→3,1→4)-β-glucanases in germinated barley

Expression sites of genes encoding (13,14)--glucan 4-glucanohydrolase (EC 3.2.1.73) have been mapped in germinated barley grains (Hordeum vulgare L.) by hybridization histochemistry. A³²P-labelled cDNA (copy DNA) probe was hybridized to cryosections of intact barley grains to localize complementary mRNAs. No mRNA encoding (13,14)--glucanase is detected in ungerminated grain. Expression of (13,14)--glucanase genes is first detected in the scutellum after 1 d and is confined to the epithelial layer. At this stage, no expression is apparent in the aleurone. After 2 d, levels of (13,14)--glucanase mRNA decrease in the scutellar epithelium but increase in the aleurone. In the aleurone layer, induction of (13,14)--glucanase gene expression, as measured by mRNA accumulation, progresses from the proximal to distal end of the grain as a front moving away from, and parallel to, the face of the scutellum.Abbreviations cDNA copy DNA - RNase ribonuclease