Differential effects of metal-binding agents on the D-glucose-6-phosphate phosphohydrolase activity of the rat liver.

The metal-binding agents (citrate, oxalate, bicarbonate, EDTA) exert dual effects on D-glucose-6-phosphate phosphohydrolase activity in the homogenate as well as in the subcellular fractions. The important differences of the effects are associated with the concentration of the chelator and with time of its addition. The small (appropriate) concentrations of the metal-binding agents stimulate and stabilize the enzyme activity. However, chelators used in higher concentrations exert the inhibitory influence on the activity of glucose-6-phosphatase. Stimulation of the reaction was observed only if the chelator was added before the enzyme-substrate complex formation has been started. The formation of the ternary complex: the enzyme(metal)-chelator substrate exerting a protective influence on the active centre has been suggested. The hypothesis of a similar action of the metal-binding agents and Pi on the glucose-6-phosphatase as a metaloproteid has been proposed.     

UDP-galactose 4-epimerase. Phosphorus-31 nuclear magnetic resonance analysis of NAD+ and NADH bound at the active site

The phosphorus atoms of NAD+ bound within the active site of UDP-galactose 4-epimerase from Escherichia coli exhibit two NMR signals, one at delta = -9.60 +/- 0.05 ppm and one at delta = -12.15 +/- 0.01 ppm (mean +/- standard deviation of four experiments) relative to 85% H3PO4 as an external standard. Titration of epimerase.NAD+ with UMP causes a UMP-dependent alteration in the chemical shifts of the resulting exchange-averaged spectra, which extrapolate to delta = -10.51 ppm and delta = -11.06 ppm, respectively, for the fully liganded enzyme, with an interconversion rate between epimerase.NAD+ and epimerase.NAD+.UMP of at least 490 s-1. Conversely, the binding of 8-anilinonaphthalene-1-sulfonate, which is competitive with UMP, causes a significant sharpening of the epimerase.NAD+ resonances but very little alteration in their chemical shifts, to delta = -9.38 ppm and delta = -12.16 ppm, respectively. UMP-dependent reductive inactivation by glucose results in the convergence of the two resonances into a single signal of delta = -10.57 ppm, with an off-rate constant for UMP dissociation from the epimerase.NADH.UMP complex estimated at 8 s-1. Reductive inactivation by borohydride under anaerobic conditions yields a single, broad resonance centered at about delta = -10.2 ppm. The data are consistent with, and may reflect, the activation of NAD+ via a protein conformational change, which is known from chemical studies to be driven by uridine nucleotide binding. Incubation of epimerase.NAD+ with UMP in the absence of additional reducing agents causes a very slow reductive inactivation of the enzyme with an apparent pseudo-first-order rate constant of 0.013 +/- 0.001 h-1, which appears to be associated with liberation of inorganic phosphate from UMP.     

Buoyancy regulation in light-limited continuous cultures of Microcystis aeruginosa

Microcystis aeruginosa was grown in light-limited continuouscultures at different growth rates on light-dark cycles at variousphotopenods. Due to the strength of the gas vesicles the organismwas not able to collapse its gas vesicles by turgor pressure.Below the maximal growth rate, the organism was buoyant dueto its high gas vesicle content. The results suggested thatthe rate of gas vesicle synthesis was not regulated. Upon atransition to high irradiance it took several hours before thecells lost their buoyancy due to polyglucan accumulation. Theresults are interpreted in an ecological context and it is suggestedthat Microcystis is an epilimnetic species due to its buoyancyregulation.     

Chemical modification of cytochrome b5, cytochrome c and myoglobin with diethylpyrocarbonate

Cytochrome b5 is required for the cytochrome P-450 LM2 catalyzed oxidation of the anesthetic methoxyflurane. The ability of cytochrome b5 to support methoxyfluorane oxidation is affected by treatment with diethylpyrocarbonate, a reagent that at neutral pH is relatively specific for histidine residues. This inactivation of cytochrome b5 is reversed with hydroxylamine, which also suggests but does not prove histidine involvement. The studies reported in this paper were undertaken to determine whether histidine modification was involved in the decrease in effectiveness of cytochrome b5, or whether the inactivation could be attributed to modification of another amino acid. Our experiments demonstrate that diethylpyrocarbonate inactivates detergent-solubilized cytochrome b5 by modifying the axial histidines and displacing the heme. Because of the unexpected ease with which diethylpyrocarbonate displaced the heme from cytochrome b5, this same process was investigated in two other hemoproteins, cytochrome c and myoglobin. Diethylpyrocarbonate could not dissociate the heme from cytochrome c, whereas the heme was lost from myoglobin even more readily than from cytochrome b5.     

Seasonal Bacterial Production in a Dimictic Lake as Measured by Increases in Cell Numbers and Thymidine Incorporation

Rates of primary and bacterial production in Little Crooked Lake were calculated from the rates of incorporation of H¹⁴CO₃⁻ and [methyl-³H]thymidine, respectively. Growth rates of bacteria in diluted natural samples were determined for epilimnetic and metalimnetic bacterial populations during the summers of 1982 and 1983. Exponential growth was observed in these diluted samples, with increases in cell numbers of 30 to 250%. No lag was observed in bacterial growth in 14 of 16 experiments. Correlation of bacterial growth rates to corresponding rates of thymidine incorporation by natural samples produced a conversion factor of 2.2 × 10¹⁸ cells produced per mole of thymidine incorporated. The mass of the average bacterial cell in the lake was 1.40 × 10⁻¹⁴ ± 0.05 × 10⁻¹⁴ g of C cell⁻¹. Doubling times of natural bacteria calculated from thymidine incorporation rates and in situ cell numbers ranged from 0.35 to 12.00 days (median, 1.50 days). Bacterial production amounted to 66.7 g of C m⁻² from April through September, accounting for 29.4% of total (primary plus bacterial) production during this period. The vertical and seasonal distribution of bacterial production in Little Crooked Lake was strongly influenced by the distribution of primary production. From April through September 1983, the depth of maximum bacterial production rates in the water column was related to the depth of high rates of primary production. On a seasonal basis, primary production increased steadily from May through September, and bacterial production increased from May through August and then decreased in September.     

Concordance of experimentally mapped or predicted Z-DNA sites with positions of selected alternating purine-pyrimidine tracts.

The recent electronmicroscopic and biochemical mapping of Z-DNA sites in phi X174, SV40, pBR322 and PM2 DNAs has been used to determine two sets of criteria for identification of potential Z-DNA sequences in natural DNA genomes. The prediction of potential Z-DNA tracts and corresponding statistical analysis of their occurrence have been made on a sample of 14 DNA genomes. Alternating purine and pyrimidine tracts longer than 5 base pairs in length and their clusters (quasi alternating fragments) in the 14 genomes studied are under-represented compared to the expectation from corresponding random sequences. The fragments [d(G X C)]n and [d(C X G)]n (n greater than or equal to 3) in general do not occur in circular DNA genomes and are under-represented in the linear DNAs of phages lambda and T7, whereas in linear genomes of adenoviruses they are strongly over-represented. With minor exceptions, potential Z-DNA sites are also under-represented compared to random sequences. In the 14 genomes studied, predicted Z-DNA tracts occur in non-coding as well as in protein coding regions. The predicted Z-DNA sites in phi X174, SV40, pBR322 and PM2 correspond well with those mapped experimentally. A complete listing together with a compact graphical representation of alternating purine-pyrimidine fragments and their Z-forming potential are presented.     

Phylogeny of Greya (Lepidoptera: Prodoxidae), based on nucleotide sequence variation in mitochondrial cytochrome oxidase I and II: congruence with morphological data

The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from nucleotide sequence variation across a 765-bp region in the cytochrome oxidase I and II genes of the mitochondrial genome. Most parsimonious relationships of 25 haplotypes from 16 Greya species and two outgroup genera (Tetragma and Prodoxus) showed substantial congruence with the species relationships indicated by morphological variation. Differences between mitochondrial and morphological trees were found primarily in the positions of two species, G. variabilis and G. pectinifera, and in the branching order of the three major species groups in the genus. Conflicts between the data sets were examined by comparing levels of homoplasy in characters supporting alternative hypotheses. The phylogeny of Greya species suggests that host-plant association at the family level and larval feeding mode are conservative characters. Transition/transversion ratios estimated by reconstruction of nucleotide substitutions on the phylogeny had a range of 2.0-9.3, when different subsets of the phylogeny were used. The decline of this ratio with the increase in maximum sequence divergence among taxa indicates that transitions are masked by transversions along deeper internodes or long branches of the phylogeny. Among transitions, substitutions of A-->G and T-->C outnumbered their reciprocal substitutions by 2-6 times, presumably because of the approximately 4:1 (77%) A+T-bias in nucleotide base composition. Of all transversions, 73%-80% were A<-->T substitutions, 85% of which occurred at third positions of codons; these estimates did not decrease with an increase in maximum sequence divergence of taxa included in the analysis. The high frequency of A<-->T substitutions is either a reflection or an explanation of the 92% A+T bias at third codon positions.      

Proposal for adoption of the base of the Undulograptus austrodentatus Biozone as a global Ordovician stage and series boundary level

The base of the Undulograptus austrodentatus Biozone appears to be a synchronous event that is widely recognizable within graptolitic facies around the world. It occurs within an interval in which graptolite species ranges are now well known and in which there is a rapid turnover in the composition of graptolite faunas. This turnover reflects the rapid evolutionary radiation of the Diplograptacea simultaneously with the appearance of several distinctive pseudisograptid and glossograptid species. These events provide the basis for the recognition of two thin but widely applicable subzones; a lower Arienigraptus zhejiangensis Subzone and an upper U. sinicus Subzone. The occurrence of the lower boundary of the U. austrodentatus Biozone within a succession of first appearances also permits accurate and reliable identification of the boundary as well as assessment of stratigraphic completeness across the boundary interval in correlated sections. Diverse graptolite faunas of late Yapeenian and early Darriwilian age occur in association with the Histiodella altifrons Biozone of the North American midcontinent conodont zonation and the Paroistodus originalis and Microzarkodina parva biozones of the North Atlantic conodont zonation. They also occur in association with the shelly-fossil zonations developed for several different continents. These features of the base of the U. austrodentatus Biozone make it a suitable level for use as the boundary level for a global stage. Its stratigraphic position within the Ordovician System relative to other likely global stages as well as its coincidence with one of the major events in graptolite evolutionary history suggest that this level also may be a suitable level for the base of a global Middle Ordovician Series.Ordovician System, Ordovician stages, graptolite zonation, chronostratigraphy, international correlation. Charles E. Mitchell and Jörg Maletz, Department of Geology, State University of New York at Buffalo, Buffalo, New York 14260-1550, USA; 13th July, 1994; revised 22nd May, 1995.     

Effect of light intensity on macromolecular synthesis in cyanobacteria

The light-dependent incorporation of NaH¹⁴CO₃ into low molecular weight compounds, polysaccharide, or protein was determined in cultures of the cyanobacteriumMerismopedia tenuissima incubated at a series of light intensities. There was an inverse relationship between incorporation into polysaccharide and protein. At light intensities of 90 E/m²/sec or above, relative incorporation of radioisotope into polysaccharide was greatest and relative incorporation into protein was lowest. Optimal relative protein accumulation occurred in samples incubated at 20 E/m²/sec. A broader optimum of light intensity for maximal protein accumulation was found if ammonia rather than nitrate was the nitrogen source. Physiological adaptation of cultures to growth at a particular light intensity did not alter the pattern of macromolecular incorporation when those cultures were tested over the series of light intensities. The response of cultures ofOscillatoria rubescens to light intensity was similar to that ofM. tenuissima, although incorporation into low molecular weight compounds was significantly greater.The effect of light intensity on macromolecular synthesis in a natural population ofOscillatoria rubescens was also determined. A pattern similar to that observed in batch cultures ofO. rubescens was occasionally found, but in other experiments there was no increase in relative protein incorporation when light intensity was decreased.