Reaction of bovine-liver copper-zinc superoxide dismutase with hydrogen peroxide. Evidence for reaction with H2O2 and HO2- leading to loss of copper

The reaction of hydrogen peroxide with the copper-zinc bovine-liver superoxide dismutase at low molar ratios (0.2-20.0) of H2O2/active site between pH 7.3-10.0 leads to the loss of native enzyme as a distinct form monitored by electrophoresis. The pH dependence of the loss of native enzyme between 7.3 and 9.0 indicates the involvement of a conjugate base on the enzyme of pKa of 8.7 +/- 0.1. The rate of loss of the native enzyme is first order with respect to the concentration of both enzyme and hydrogen peroxide between pH 7.3 and 9.0 with no evidence for binding of peroxide. A second-order rate constant of 3.0 +/- 1.0 M-1 s-1 is obtained from these data. At pH 10.0 the reaction is first order with respect to enzyme concentration but saturable in H2O2. All data are consistent with the interpretation that H2O2 reacts with the enzyme at the lower pH where the reaction is dependent upon the conjugate base of a functional group on the enzyme. At the higher pH, the data are consistent with the reaction of HO2- and H2O2 with the dismutase. The dissociation constant for HO2- calculated from the kinetic data at pH 10.0 is between 25-50 microM and the rate constant for the breakdown of the HO2- dismutase complex is 1.10 + 0.05 x 10(-2) s-1. The change in the electrophoretic pattern at all pH values is accompanied by the loss of the ability of the enzyme to bind copper. Weakly bound or free copper can be detected using bathocuproine disulfonate. Furthermore copper-defficient forms of the enzyme can be detected by staining gels of the peroxide-treated dismutase with diethyldithiocarbamate.     

Intrinsic potential for high fetal hemoglobin production in a Druze family with β-thalassemia is due to an unlinked genetic determinant

Summary The mechanism for elevated production of fetal hemoglobin (Hb F) in a Druze patient with °-thalassemia intermedia was investigated. Heterozygous family members exhibited normal Hb F levels, suggesting that the increase in -gene expression in the propositus may be partly due to anemic stress. Erythroid progenitors of these family members cultured in vitro [burst forming units (erythroid); (BFUe)] showed elevated synthesis of Hb F, indicating the existence of a genetically determined intrinsic capacity for high Hb F production in this family. The propositus was found to be homozygous for a IVS2-position 1 mutation, on the background of Mediterranean haplotype I, which is not known to be linked to high Hb F production. Moreover, extensive molecular studies of the -globin gene cluster, including sequence analysis of the promoter regions of the -globin genes, did not reveal any cisacting mechanism that could account for the high Hb F production in the propositus. A young niece of the propositus with °-thalassemia major was recently discovered, who was homozygous for the same -globin allele and haplotype as the propositus. However, unlike her uncle, she does not have a high Hb F level and presents with a severe clinical course. Her inability to produce high Hb F suggests that the genetic determinant for increased -gene expression in the propositus is unlinked to the -globin gene cluster.     

The critical region on the human Xq

Summary Adult female carriers of balanced X; autosome translocations (118 cases) and of balanced X inversions (31 cases) have been collected from the literature. Forty-five of the 118 translocation carriers in whom the break was in the critical region (Xq13–q22, Xq22–q26, separated by a narrow region within Xq22) showed gonadal dysgenesis. Seven of the 31 inversion carriers in whom the break was in the same region also had gonadal dysgenesis, whereas the remaining 24 were normal in this respect. The critical region consists mainly of Q-bright material, and is the fifth brightest segment in the human genome. The region contains relatively few genes. It is possible that meiotic crossing-over, rarely, if ever, takes place in it. The critical region may therefore consist of two supergenes whose integrity must be maintained to allow normal ovarian development. The effect exerted by this region differs from other known position effects, in that it is independent of the break-point within the region and of the chromosome bands to which the broken ends are attached. One possible mechanism causing this effect might be a change in the replication order of the chromosome bands, which, in turn, might affect their function.     

Enhanced tumor susceptibility of immunocompetent mice infected with lymphocytic choriomeningitis virus

Summary Mice infected i.v. with high doses of lymphocytic choriomeningitis virus (LCMV; 10⁵–10⁶ plaqueforming units) 8–10 days prior to challenge with the methylcholanthrene-induced fibrosarcoma tumor cell line MC57G or the melanoma cell line B16 tumor cells showed an enhanced tumor susceptibility with respect to both growth kinetics of the tumor and the minimal dose necessary for tumor take. After transient initial growth, MC57G tumor cells were all rejected by uninfected C57BL/6 mice by day 14. Mice preinfected i.v. with LCMV 3 weeks before or at the time of tumor challenge, but not those infected 2 months before or 7 days after, showed increasing tumor growth, the tumor take being 100% for 10⁶, 50% for 10⁵ and 37% for 10⁴ MC57G tumor cells injected into the footpad compared with resistance to 10⁶ cells in normal mice. B16 melanoma cells also grew more rapidly in LCMV-preinfected mice and by day 40 tumors were established with about 100 times fewer cells, i.e. about 10³ compared with 3×10⁴–3×10⁵ for uninfected mice. Analysis of the growth of tumor cells in normal and in LCMV-carrier mice revealed that the latter mice were not more susceptible to LCMV-infected than to uninfected MC57G. Since LCMV-carrier mice fail to mount LCMV-specific T cell responses, these results suggest that anti-LCMV-specific T cells may be responsible for acquired immunodeficiency hampering immune surveillance against the tumors studied.Supported by grants from the Swiss National Science Foundation 3.259–0.87 and the Kanton of Zürich     

Meristem transformation of sunflower via Agrobacterium

Summary For transformation of sunflower (Helianthus annuus L. cv. Zebulon), shoot apical meristems were dissected from seeds and cocultivated with a disarmed Agrobacterium tumefaciens strain harboring a binary vector carrying genes encoding GUS- and NPTII-activity. The influence of the media conditions, the time of cocultivation and the stage of the developing seed on shoot development and meristem transformation was analysed. Transformants were selected by their ability to grow on kanamycin. Transformation was confirmed by assays for GUS and NPTII. GUS-positive shoots were rooted on rockwool and transferred to soil. Transformation of shoot meristem cells occurred at low frequencies. Chimaeric expression of the two genes was observed in transformed plants. Integration of the foreign DNA in the sunflower genome was confirmed with the polymerase chain reaction.Abbreviations GUS ß-Glucuronidase - NPTII Neomycin phosphotransferase II     

Wealth and pastoral dairy production: A case study from Maasailand

Much of pastoral development in Africa has been predicted on the assumed desirability of converting pastoralists to commercial beef producers. Such development has ignored two fundamental aspects of pastoral production: first, the greater human support capacity of a dual milk/meat production system, and second, significant wealth inequality within pastoral communities. This paper presents a case study based on several years of field research in Kenya Maasailand; it examines variations by wealth status in milking strategies and the level of milk offtake for human consumption. Rich households have five times the number of cattle per reference adult as poor households, but similar levels of milk consumption, due to differences in the allocation of milk between calves and people. Residential location and watering frequency also vary by wealth status, contributing to a difference in total milk production per cow.The research and the major part of the writing of this paper were done while the author was a scientist with the Kenya Country Program of the International Livestock Center for Africa. Finishing touches were done at the International Laboratory for Research on Animal Disseases. Neither organization bears responsibility for the content. I wish to thank P. N. deLeeuw for long and valuable discussions; the animal science aspects of the research also benefitted from discussions with K. Wagenaar and M. Nicholson. P. Lembuya was, as always, a faithful and demanding assistant. The useful comments of S. Sandford, T. Conelly, U. Herren, and J. Ensminger are also appreciated.     

CO2 fixation in halobacteria

Seven strains of extremely halophilic bacteria (Halobacterium spp., Halococcus spp., and Haloarcula sp.) fixed CO₂ under light and dark conditions. Light enhanced CO₂ fixation in Halobacterium halobium but inhibited it in Halobacterium volcanii and Haloarcula strain GN-1. Propionate stimulated ¹⁴CO₂ incorporation in some strains, but inhibited it in others. Semi-starvation in basal salts plus glycerol induced enhanced CO₂ fixation rates. ¹⁴CO₂ fixation in semi-starved cells was stimulated by NH ₄ ⁺ or pyruvate and inhibited by succinate and acetate in most strains. No possible reductant was found. In cell-free extracts of H. halobium, NH ₄ ⁺ but not propionate stimulated ¹⁴CO₂ fixation. No RuBP carboxylase activity was detected. The main ¹⁴C-labeled -keto acid detected after a 2-min incubation with ¹⁴CO₂ and pyruvate was pyruvate. Little or no -ketobutyrate was detected among the early products of propionate-stimulated CO₂ fixation. Glycine was the major amino acid synthesized during a 2-min incubation with NH ₄ ⁺ , propionate, and ¹⁴CO₂. Propionate-stimulated CO₂ fixation was sensitive to trimethoprim and insensitive to avidin. A novel pathway for non-reductive CO₂ fixation involving a glycine synthase reaction with CO₂, NH ₄ ⁺ , and a methyl carbon derived from the -carbon cleavage of propionate is tentatively proposed.Abbreviations used BBS buffered basal salts - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid - MOPS 3-(N-morpholino)propanesulfonic acid - DNPH 2,4-dinitrophenylhydrazine - DNP dinitrophenyl - TLC thin-layer chromatography - FH₄ tetrahydrofolate This work was supported by National Science Foundation grant PCM-8116330 and Petroleum Research Fund grant PRF 13704-AC2     

Spectral mechanisms in the tree squirrel retina

Summary The retina of the gray squirrel (Sciurus carolinensis) contains rods and cones in a ratio of about 23. The spectral mechanisms in this retina were examined in behavioral and electrophysiological experiments. Tests of color vision revealed that this animal has a spectral neutral point at about 500 nm and, thus, dichromatic color vision. Recordings made from single optic nerve fibers and results obtained from an analysis of the flicker photometric electroretinogram (ERG) indicated that vision in the gray squirrel is based on three spectral mechanisms. One of these, presumably rod-based, has peak sensitivity at about 502 nm. The other two mechanisms reflect the presence of two classes of cone having average peak sensitivity of about 444 nm and 543 nm.     

Acoustic orientation in adult, female crickets (Gryllus bimaculatus de Geer) after unilateral foreleg amputation in the larva

Summary InGryllus bimaculatus females one foreleg was amputated at the coxa-trochanter joint in the 2nd, 4th or 8th/9th larval instar. A leg of up to normal length is regenerated (Fig. 1) but it lacks a functional ear. In spite of the, usually shorter, regenerated foreleg, the adult one-eared crickets show no impairments in walking when tested on a locomotion compensator. Without sound they walk erratically and most of them weakly circle towards the intact side (Fig. 2).With calling song presentation three response types can be distinguished:tracking (Fig. 3A), hanging on (Fig. 3B) or continuouscircling towards the intact side (Fig. 3C, D). Turning tendencies in monaurals increase with song intensity and exceed those of intact and bilaterally operated animals (Fig. 4). Course deviations towards the intact side also slightly increase with intensity (Fig. 5). Course stability is reduced compared to that of intact animals but exceeds that of bilaterally operated crickets (Figs. 5, 6). It is best at 60 dB and deteriorates at higher sound intensities (Fig. 6). The percentage of monaurals tracking or hanging on decreases with increasing intensity (Fig. 7B). Tracking is established in most animals but it is limited to a narrow intensity range (Fig. 7A, C). Apart from an increased percentage of tracking after early operations (Fig. 7D), there are no prominent changes in orientational parameters with the date of foreleg amputation.Reamputation of the regenerated leg in the adult monaurals does not significantly impair acoustic orientation (Figs. 8, 9), but occlusion of the ipsilateral prothoracic spiracle does (Figs. 10, 11).An attempt is made to correlate the behavioral performance with the activity of auditory interneurons which have undergone morphological and physiological changes (Fig. 12).     

Structural relationships among chloramphenicol-resistance plasmids of Staphylococcus aureus

Abstract Twelve different chloramphenicol-resistance (Cm^r) plasmids detected in Staphylococcus aureus strains isolated between 1952 and 1981 were characterized by restriction endonuclease, DNA hybridization and heteroduplex analyses. These studies revealed three families of Cm^r plasmids which were distinguished by their chloramphenicol acetyltransferase sequences; the prototype plasmids of the families were pC221, pC223 and pC194. The cat and replication ( rep ) genes of the plasmid pC221 were highly conserved in other pC221 family members and were related to their homologs in the pC223 family plasmids; however, the cat and rep genes of the pC194 family plasmids were distinct.