Computer-assisted mathematical analysis of sigmoid biological events

A program in BASIC is described which allows accurate quantificationof some numerical parameters that can be objectively correlatedto biological indexes in sigmoid biological events. Attentionwas focused on the polymerization process of actin (a muscleprotein with a mol. wt of 42 000 daltons) studied as the variationin the OD₃₆₀ index with time. The experimental points, if plotted,can be well approximated by a rational function of the typeOD₃₆₀ = f(t), which passes through the origin and can be representedgraphically by a sigmoid curve. The program was very helpfulin comparing the experimental curves and in analysing significantparameters, such as maximum velocity and asymptote, that characterizethese curves and whose interpretation would otherwise be purelysubjective. Received on July 11, 1985; accepted on January 13, 1986     

Metabolism and Solubilization of Cellulose by Clostridium cellulolyticum H10

When Clostridium cellulolyticum was grown with cellulose MN300 as the substrate, the rates of growth and metabolite production were found to be lower than those observed with soluble sugars as the substrate. At low cellulose concentrations, the growth yields were equal to those obtained with cellobiose. The main fermentation products from cellulose and soluble sugars were the same. Up to 15 mM of consumed hexose, a change in the metabolic pathway favoring lactate production similar to that observed with soluble sugars was found to occur concomitantly with a decrease in molar growth yield. With cellulose concentrations above 5 g/liter, accumulation of soluble sugars occurred once growth had ceased. Glucose accounted for 30% of these sugars. A kinetic analysis of cellulose solubilization revealed that cellulolysis by C. cellulolyticum involved three stages whatever cellulose concentration was used. Analysis of these kinetics showed three consecutive enzymatic activity levels having the same K_m (0.8 g of cellulose per liter, i.e., 5 mM hexose equivalent) but decreasing values of V_max. The hypothesis is suggested that each step corresponds to differences in cellulose structure.     

Localization of alpha 1-microglobulin (HC protein) in normal human tissues: an immunohistochemical study using monoclonal antibodies

Summary Alpha-1-microglobulin is a low molecular weight (approximately 30 000 d) glycoprotein present in biological fluids. It is heterogeneous in charge. A monoclonal antibody was used to investigate the tissue distribution of the protein in normal human tissues and cell lines by indirect immunofluorescence and immunoperoxidase techniques. The protein was demonstrable in cells of the monocyte-macrophage lineage, in thymus and T cell dependent areas of spleen, lymph node and tonsils. It was detected in several lymphoid or nonlymphoid cell lines but not in peripheral blood lymphocytes. The microglobulin was also detectable in the cytoplasm of hepatocytes. Finally, it was observed in glandular secretions (sudoral glands and mucosal glands of the digestive tract) where it may be associated with IgA. Possible explanations for the highly divergent results previously reported with polyclonal antisera to ₁ microglobulin are discussed.     

An acetylenic mechanism-based inhibitor of dopamine beta-hydroxylase

The catalytic action of dopamine beta-hydroxylase on 1-phenyl-1-propyne results in concomitant loss of enzyme activity. At pH 5.5 and 25 degrees C, 1-phenyl-1-propyne inactivates dopamine beta-hydroxylase in a mechanism-based fashion. The inactivation rate is first-order, follows saturation kinetics, and is strictly dependent on catalysis (oxygen and ascorbate are essential). The inactivation rate of saturating 1-phenyl-1-propyne (kinact) increases from 0.08 to 0.22 min-1 when the oxygen saturation increases from 21 to 100%, respectively. Inactivation also requires a copper-containing catalytically competent enzyme. Tyramine and norepinephrine (respectively, substrate and product of the normal catalytic reaction) protect against inactivation, and no regain of enzyme activity occurs after prolonged dialysis. Experiments with ether-extracted incubation solutions (+/- enzyme) showed no difference in their gas chromatography-mass spectral patterns implying that inactivation of dopamine beta-hydroxylase by 1-phenyl-1-propyne occurs through a kinetic process with a partition ratio (kcat/kinact) equal to or near 1. Thus, this acetylenic substrate analog appears to be a very efficient mechanism-based inhibitor of dopamine beta-hydroxylase. We propose that inactivation of this enzyme by 1-phenyl-1-propyne proceeds by formation of a reactive intermediate that occurs prior to product formation and that alkylates an amino acid residue at the active site of the enzyme.     

Effects of temperature and CO2 enrichment on kinetic properties of phospho-enol-pyruvate carboxylase in two ecotypes of Echinochloa crus-galli (L.) Beauv., a C4 weed grass species

Summary Two populations of Echinochloa crus-galli (Québec, Mississippi) were grown at the Duke University Phytotron under 2 thermoperiods (28°/22°C, 21°/15°C day/night) and 2 CO₂ regimes (350 and 675 l l^-1). Thermostability, energy of activation (E _a ),K _m (PEP), K _m (Mg⁺⁺), and specific activity of phospho-enol-pyruvate carboxylase (PEP_c) were analyzed in partially purified enzyme preparations of plants grown for 5 weeks. Thermostability of PEP_c from extracts (in vitro) and leaves (in situ) was significantly higher in Mississippi plants. In vitro denaturation was not appreciably modified by thermal acclimation but CO₂ enrichment elicited higher thermostability of PEP_c. In situ thermostability was significantly higher than that of in vitro assays and was higher in Mississippi plants acclimated at 28°/22°C and in plants of the two ecotypes grown at 675 l l^-1 CO₂. E _a (Q ₁₀ 30°/20°C) for PEP_c was significantly lower in Québec plants as compared to Mississippi and no acclimatory shifts were observed. Significantly higher K _m's (PEP) in 20°C assays were obtained for Mississippi as compared to Québec plants but values were similar at 30°C and 40°C assays. K _m (Mg⁺⁺) decreased at higher assay temperatures and were significantly lower for PEP_c of the Québec ecotype. No significant changes in K _m (Mg⁺⁺) values were associated with modifications in temperature on CO₂ regimes. PEP_c activity measured at 30°C was significantly higher for Québec plants when measured on a leaf fresh weight, leaf area or protein basis but not on a chlorophyll basis. Significantly higher PEP_c activity for both genotypes was observed for plants acclimated at 21°/15°C or grown at 675 l l^-1 CO₂. Net photosynthesis (Ps) and net assimilation rates (NAR) were higher in Québec plants and were enhanced by CO₂ enrichment. NAR was higher in plants acclimated at low temperature, while an opposite trend was observed for Ps. PEP_c activities were always in excess of the amounts required to support observed rates of CO₂ assimilation.     

Nucleic acid hybridisation with a probe specific for 3 aminoglycoside acetyltransferase type IV: A survey of resistance to apramycin and gentamicin in animal strains of Escherichia coli

Abstract Resistance to apramycin due to production of a 3-aminoglycoside acetyltransferase type IV (AAC(3)IV) has recently been detected among Gram-negative bacteria isolated in France from bovine clinical samples. 24 apramycin-resistant Escherichia coli strains isolated over the country, and epidemiologically unrelated, were studied by colony hybridization using an intragenic probe specific for AAC(3)IV. The results obtained indicated that the structural gene for the acetyltransferase was present in all the isolates tested and in the corresponding apramycin-resistant transconjugants. This observation demonstrates that resistance to apramycin by acetylation of the antibiotic has spread very rapidly in bovine Gram-negative bacteria.     

Glutamate synthase in rice roots. Studies on the electron donor specificity

Rice root glutamate synthase activity was assayed with various reducing systems. Ferredoxin-dependent glutamate synthase (EC 1.4.7.1) and pyridine nucleotide-dependent glutamate synthase (NADH, EC 1.4.1.14; or NADPH, EC 1.4.1.13) exhibited a strict specificity for the electron donor. The ferredoxin-dependent glutamate synthase from rice roots could accept electrons from photoreduced ferredoxin in an illuminated reconstituted spinach chloroplast system. Thioredoxin, a potent electron carrier, was not able to provide either ferredoxin-dependent or pyridine nucleotide-dependent glutamate synthase with electrons as no glutamate formation was detected in the presence of reduced thioredoxin f or m.