Regulation of induction of phenylalanine ammonia-lyase in suspension cultures of Phaseolus vulgaris

Total RNA was extracted from fast growing suspension cells of bean, the mRNA was translated and the products of protein synthesis analysed by gel electrophoresis. Actinomycin D (20 g ml^–1) added to the cultures 12 h before the induction of phenylalanine ammonia-lyase (PAL) activity by naphthylacetic acid (NAA) (1 mg/l) and kinetin (0.2 mg/l) failed to prevent the increased activity of the enzyme usually produced by this ratio of the plant growth hormones. PAL was isolated and purified from suspension cultured bean cells. The purified enzyme ran as a single band on polyacrylamide gel electrophoresis. The protein translated from RNA prepared from induced and non-induced cells was separated by gel electrophoresis and the bands of protein on the gels were compared. There was no evidence for an increase in the amount of PAL synthesised in vitro from the mRNA of induced cells even though these had 5 times the amount of activity of the enzyme compared with that of the non-induced cells. The results indicate that the induction of PAL activity is not immediately preceeded by an increase in the synthesis of PAL-mRNA by the cells. The control of the activity of the enzyme is discussed with respect to this finding.Abbreviations PAL phenylalanine ammonia-lyase - NAA 3naphthylacetic acid - DEAE Diethylamino ethyl - EDTA Ethylenediamine tetraacetate - SDS Sodium dodecyl sulphate     

Metal tolerance in tissue cultures of anthoxanthum odoratum

Tissue cultures were initiated from one non-tolerant (S20) and two zinc and lead tolerant (T92 and T94) clones of Anthoxanthum odoratum. Growth of callus from the non-tolerant clone was reduced by the presence of zinc, lead, copper and nickel, whereas callus from the two tolerant clones showed no reduction of growth in the presence of zinc and lead but growth was reduced by copper and nickel. The specificity of metal tolerance shown by the parental material was maintained in the callus. Tolerant and non-tolerant callus accumulated similar amounts of zinc and lead.     

The role of serine hydroxymethyltransferase in cell proliferation: DNA synthesis from serine following mitogenic stimulation of lymphocytes

It is shown that the time-course of incorporation of radioactivity from [3-¹⁴C]serine into nucleic acids parallels DNA synthesis following mitogenic stimulation of human peripheral blood lymphocytes by phytohaemagglutinin (PHA). The activity of serine hydroxymethyltransferase was elevated about four-fold in PHA-stimulated lymphocytes compared to that in unstimulated control ceils. It is suggested that lymphocytes, in common with other proliferating cell systems:, may synthesize serine de novo for utilization in pathways of nucleotide biosynthesis following mitogenic stim--ulation.     

The learning skills of primates: The rhesus macaque in comparative perspective

Twenty-month-old rhesus monkeys were tested in a modified discrimination-reversal paradigm, which was designed to distinguish abstract learning from stimulus-response associational learning. Previous studies indicate that talapoin monkeys learn associationally and great apes via forming abstract concepts. Adult rhesus monkeys are apparently capable of forming simple abstractions, but learn primarily through associational process. The results of this study show the adolescent rhesus monkeys to be associational learners, with their response patterns indicating more complexity than the talapoins but less than the adult rhesus monkeys. The data suggest that rhesus monkeys develop their low-level capacity of abstract learning with maturation.     

Evolution of enzyme catalytic power. Characteristics of optimal catalysis evaluated for the simplest plausible kinetic model

1. Evolutionary changes in the structure of an enzyme that provide an increase in its K_m value are considered. Provided that K_m increases as a result of increases in the forward rate constants of the catalysis relative to the reverse rate constants, the enzyme catalyses the conversion of a fixed concentration of its substrate more rapidly when its structure provides that K_m>[S] than when K_m<[S]. 2. Catalytic efficiency of enzymes is discussed in terms of the simplest plausible model, the Haldane [(1930) Enzymes, Longmans, London] reversible three-step model: [Formula: see text] The rate equation for the forward reaction of this model (formation of P) may be written in the simple form: [Formula: see text] K_eq. is the equilibrium constant (=[P]_eq./[S]_eq.), and k_cat.=V/[E]_T, where [E]_T is the total enzyme concentration. 3. To assess the effectiveness of an enzyme, it is necessary only to determine the extent to which the constraints of a particular kinetic mechanism permit v₂ (v when K_m»[S]) to approach v_d (the diffusion-limited rate). 4. The value of the optimal rate of catalysis (v_opt., the maximal value of v₂) is dictated by the equilibrium constant for the reaction, K_eq.; v₂=v_d/a, where [Formula: see text] when k₊₁ is assumed equal to k₋₃, and v_opt.=v_d/a_min.. When K_eq.≥1, it is necessary that k₊₂»k₋₁ for a to take its minimum value, a_min.; when K_eq.«1, it is necessary only that k₊₂»K_eq.·k₋₁, i.e. a can equal a_min. even if k₊₂<k₋₁. When K_eq.»1, v_opt.=v_d; when K_eq.=1, v_opt.=v_d/2, and when K_eq.«1, v_opt.=K_eq.·v_d. 5. The analysis, together with predicted effects of evolutionary pressure, suggests that in practice the rates of the fastest enzyme-catalysed freely reversible reactions might be expected to be lower than the value of k₊₁[E]_T[S] by about an order of magnitude, particularly if K_eq.<1. 6. The existing literature suggests that, in general, appropriate values of K_m have evolved for the provision of high rates of catalysis but that many values of k_cat. are not large enough to provide optimal rates of catalysis unless the value of k₊₁ in vivo is lower than its value in free solution.     

Preparation and characterization of isolated parenchymal cells from guinea pig liver

1. A method is described for the preparation of isolated cells from guinea pig liver. This involved perfusion in situ, in the non-physiological direction, with collagenase. 2. The cell yield was 20--30%, comparable with those from the livers of other species. 3. The ratio of lactate dehydrogenase to glutamate dehydrogenase in the cells was similar to that in vivo, indicating that there was negligible leakage of cytoplasmic enzymes. 4. The concentrations of K+ and adenine nucleotides were initially lower than in the perfused liver; normal values were obtained on incubation, particularly in the presence of substrate. 5. The L-lactate: pyruvate ratio is 16:1, close to established values. The total beta-hydroxybutyrate: acetoacetate ratio indicates that the mitochondrial redox state is more oxidised than in the perfused liver, but the intracellular ratio is similar to that of the intact liver. 6. Rates of gluconeogenesis and ureogenesis, are within the physiological range. Maximal gluconeogeneis from L-lactate was preceded by a lag period. L-lysine stimulated glucose production from L-lactate but did not abolish the lag phase. 7. The effects of aminooxyacetate and octanoate on L-lactate gluconeogenesis were similar to those in the perfused liver.     

Common Sequence at the 5′ Ends of the Segmented RNA Genomes of Influenza A and B Viruses

Guanylyl- and methyltransferases, isolated from purified vaccinia virus, were used to specifically label the 5′ ends of the genome RNAs of influenza A and B viruses. All eight segments were labeled with [α-³²P]guanosine 5′-triphosphate or S-adenosyl[methyl-³H]methionine to form “cap” structures of the type m⁷G(5′)pppN^m-, of which unmethylated (p)ppN- represents the original 5′ end. Further analyses indicated that m⁷G(5′)pppA^m, m⁷G(5′)pppA^mpGp, and m⁷G(5′)pppA^mpGpUp were released from total and individual labeled RNA segments by digestion with nuclease P1, RNase T1, and RNase A, respectively. Consequently, the 5′-terminal sequences of most or all individual genome RNAs of influenza A and B viruses were deduced to be (p)ppApGpUp. The presence of identical sequences at the ends of RNA segments of both types of influenza viruses indicates that they have been specifically conserved during evolution.     

Seed glycoproteins of Lupinus angustifolius

The seed globulins of Lupinus angustifolius are glycoproteins containing 1.4–1.9% (α-conglutin), 2.8–6.4 % (β-conglutin) and 1.2–3.8% (γ-conglutin) carbohydrate. The highest values were obtained after acid hydrolysis and determination by phenol—H₂SO₄, (α, γ-conglutins) or by methanolysis and sugar determination by GLC (β-conglutin). TCA denaturation of β- and γ-conglutins was necessary to remove adsorbed galactomannans before determination of glycoprotein carbohydrates. All 3 conglutins contained mannose, galactose and glucosamine, though the ratio of mannose to galactose, and to a lesser extent neutral sugars to hexosamine varied. Small amounts of fucose were found associated only with γ-conglutin.     

The effects of cholesterol on the time-resolved emission anisotropy of 12-(9-anthroyloxy)stearic acid in dipalmitoylphosphatidylcholine bilayers

The time-resolved fluorescence emission anisotropy of 12-(9-anthroyloxy)stearic acid (12-AS) and 1,6-diphenyl-1,3,5-hexatriene (DPH) have been measured in dipalmitoylphosphatidylcholine liposomes in the presence and absence of 40 mol% cholesterol at temperatures above and below the phase transition temperature (41°C). By using a synchronously-pumped mode-locked frequency-doubled dye laser and single photon counting detection with an excitation response function of 300 picosecond, rotational correlation times down to less than 1 nanosecond could be resolved. Whereas DPH showed only small changes in the limiting anisotropy on the addition of cholesterol, 12-AS showed significant increases in this parameter with the effect being potentiated at higher temperatures. This difference in behaviour has been attributed to a fluorophore-cholesterol interaction that resulted in a change in the fluorophore geometry. Not only do DPH and 12-AS sense different depolarizing rotations due to the different directions of their emission dipoles but also differ in their lipid interactions which alter their limiting anisotropies. The implication is that the comparison of steady-state anisotropy measurements between chemically identical fluorophores in different lipid environments may be complicated by molecular distortions that change the motions to which the steady-state fluorescence parameters will be sensitive.