Metabolism of a phenylcoumaran substructure lignin model compound in ligninolytic cultures of Phanerochaete chrysosporium

The degradation of the phenylcoumaran substructure model compound methyl dehydrodiconiferyl alcohol by the white-rot wood decay fungus Phanerochaete chrysosporium was investigated using culture conditions optimized for lignin oxidation. Initial attack was in the cinnamyl alcohol side chain, which was oxidized to a glycerol structure. This was subsequently converted by loss of the two terminal carbon atoms, C and C, to yield a C-aldehyde structure, which was further oxidized to the C-acid compound. The next detected intermediate, a phenylcoumarone, was produced by double bond formation between C and C, and oxidation of the C-alcohol to an aldehyde group. Further oxidation of C to an acid yielded the next intermediate. The final identified degradation product was veratric acid. No products from the 5-substituted aromatic ring, and no phenolic products, were found. The initial glycerol-containing intermediate was a mixture of the threo and erythro forms, and no optical activity could be found, suggesting that its formation might have involved nonstereospecific C-C epoxidation followed by non-enzymatic hydrolysis of the epoxide.Abbreviations TLC thin layer chromatography - LDA lithium diisopropyl amide - DDQ 2,3-dichloro-5,6-dicyanobenzoquinone - MS mass spectrometry - UV ultraviolet spectroscopy     

Kinetics of growth and lactic acid production in continuous and batch culture

Summary In a lactic acid fermentation by Streptococcus faecalis, the specific consumption rate of glucose (v) and the specific production rate of lactic acid () were represented by the following simple equations as functions of the specific growth rate (): 1/=(1/) + 1/ = (1/) + By use of data from a batch culture, these two equations were derived from enzyme kinetics of the product inhibition. These equations were successfully applied to the results of batch culture and chemostat culture. In addition, calculation of ATP yield by these equations agreed with the experimental results better than the conventional Leudeking-Piret type equation, which includes two terms associated with growth and not with growth. Correspondence to: H. Ohara     

Carbon-dioxide exchange in lichens: determination of transport and carboxylation characteristics

Measurements were made of net rates of CO₂ assimilation in lichens at various ambient concentrations of CO₂ in air and in helox (79% He, 21% O₂). Because of the faster rate of CO₂ diffusion in the pores of lichen thalli when filled with helox than when filled with air, a given net rate of assimilation was achieved at a lower ambient concentration of CO₂ in helox. The differences were used to estimate resistances to diffusion through the gas-filled pore systems in lichens. The technique was first tested with five lichen species, and then applied in a detailed study with Ramalina maciformis, in which gas-phase resistances were determined in samples at four different states of hydration and with two irradiances. By assuming, on the basis of previous evidence, that the phycobiont in R. maciformis is fully turgid and photosynthetically competent at the smallest hydration imposed (equilibration with vapour at 97% relative humidity), and that, with this state of hydration, diffusion of CO₂ to the phycobiont takes place through continuously gas-filled pores, it was possible also to determine both the dependence of net rate of assimilation in the phycobiont on local concentration of CO₂ in the algal layer, and, with the wetter samples, the extents to which diffusion of CO₂ to the phycobiont was impeded by water films. In equilibrium with air of 97% relative humidity, the thallus water content being 0.5 g per g dry weight, the resistance to CO₂ diffusion through the thallus was about twice as large as the resistance to CO₂ uptake within the phycobiont. Total resistance to diffusion increased rapidly with increase in hydration. At a water content of 2 g per g it was about 50 times as great as the resistance to uptake within the phycobiont and more than two-thirds of it was attributable to impedance of transfer by water. The influences of water content on rate of assimilation at various irradiances are discussed. The analysis shows that the local CO₂ compensation concentration of the phycobiont in R. maciformis is close to zero, indicating that photorespiratory release of CO₂ does not take place in the alga, Trebouxia sp., under the conditions of these experiments.Symbols and Units rate of CO₂ diffusion in air relative to that in carrier gas (unity if the carrier gas is air and 0.43 if is helox) - A₁ net rate of CO₂ uptake by the lichen - A_p gross rate of carboxylation minus photorespiratory decarboxylation in the phycobiont, i.e. net rate of light-activated CO₂ exchange - A_* maximum, CO₂-saturated magnitude of A_p - c concentration of CO₂ - c_a ambient concentration of CO₂ - c_i c_a minus difference in CO₂ concentration across air-filled pore space in the thallus - c₈ CO₂ concentration equivalent to partial pressure of CO₂ at the surface of the phycobiont - ₁ magnitude of c_a at which A₁ = 0 - _* magnitude of c_* at which A_p = 0 - R rate of dark respiration in the lichen (mycobiont and phycobiont) - R rate of dark respiration in region between the surface of the lichen and an arbitrary distance from the surface within the thallus - r resistance to CO₂ transfer from lichen surface to the surface of the phycobiont - r resistance to CO₂ transfer between effective source of dark respiration in the lichen and the surface of the phycobiont - r_g, r _g components of r and r, respectively, attributable to transfer in air-phase - r_w, r _w components of r and r, respectively, attributable to transfer in water-phase - r component of r between surface of lichen and an arbitrary distance from the surface within the thallus - r_* resistance to CO₂ transfer and carboxylation in the phycobiont - RH relative humidity     

Relationships between carbon and hydrogen isotope ratios and nitrogen levels in leaves ofClusia species and two other Clusiaceae genera at various sites and different altitudes in Venezuela

Samples of the Clusiaceae generaClusia, Oedematopus andDystovomita were collected at various sites and different altitudes in northern and south-western Venezuela. Analyses of stable isotopes of carbon and hydrogen and of leaf-nitrogen levels were performed on the dried samples. Correlations among these variables, i.e. carbon isotope discrimination (), hydrogen isotope ratio (D) and N-levels, and with altitude were assessed. In the samples, where values of above 15 indicate predominant performance of C₃ photosynthesis, there were slight tendencies of increasing , D and N-levels with increasing altitude and of increasing with increasing N. Although these correlations taken separately were not statistically significant, they support each other and indicate increasing transpiration and increased leaf-nutrient supply at increasing altitude. Performance of crassulacean acid metabolism (CAM) in species ofClusia appears to be restricted to altitudes below 1500 m a.s.l. There was a significant negative correlation of with altitude in the samples, where values of below 10 indicated predominant performance of CAM. This suggests that phases II and IV of CAM are progressively suppressed towards the upper altitudinal limit of CAM inClusia in northern Venezuela. It is concluded that among the large number of environmental factors and combinations thereof, which determine the expression of CAM inClusia and trigger C₃-CAM transitions in C₃/CAM intermediate species, low availability of water is the most important.     

Steady state changes in mitochondrial electrical potential and proton gradient in perfused liver from rats fed a high fat diet

In this work the protonmotive force (p), as well as the subcellular distribution of malate, ATP, and ADP were determined in perfused liver from rats fed a low fat or high fat diet, using density gradient fractionation in non acqueous solvents.Rats fed a high fat diet, despite an enhanced hepatic oxygen consumption, exhibit similar p to that found in rats fed a low fat diet, but when we consider the two components of p, we find a significant decrease in mitochondrial/cytosolic pH difference (pH_m) and a significant increase in mitochondrial membrane potential (_m) in rats fed a high fat diet compared to rats fed a low fat diet, which tend to compensate each other. In rats fed a high fat diet the concentration ratio of malate and ATP/ADP does not reflect the changes in pH_m and _m, which represent the respective driving force for their transport.The findings are in line with an increase in substrate supply to the respiratory chain which is, however, accompanied by a higher energy turnover in livers from HFD rats. By this way the liver could contribute to the lack of weight gain from the high caloric intake in HFD rats.     

Thermocyclops ouadanei n. sp. (crustacea: copepoda) from Mauretania (North-West Africa)

Thermocyclops ouadanei n. sp. from Mauretania is described and figured. This new species is characterized by the structure of the receptaculum seminis, the uniting lamella of the P₄ which bears no marginal denticles on the rounded prominences, the length of the apical spines on the endopodite 3 of the P₄ and by the length of the furcal setae. Its relationship with other Thermocyclops species is discussed.Contribution no 16 from project Limnology of the Sahara under contract n° 2.0009.75 with the Fonds voor Kollektief Fundamenteel Onderzoek, Belgium     

Mutagenic effectiveness and efficiency of EMS,DES and gamma-rays in rice

Summary Data on chlorophyll mutation frequency after treatment with EMS, DES and gamma-rays and sequential administration of gamma-rays and the two alkylating agents in three varieties of rice have been used to work out quantitatively the effectiveness and efficiency of each mutagen and combination treatment. For effectiveness, the order is EMS > DES and for efficiency it is EMS > DES > gamma-rays. In some sequential treatments (Gamma-rays + DES in IR8 and Basmati; DES + gamma-rays in IR8 and Jhona; Gamma-rays + EMS in IR8 and Basmati; and EMS + gamma-rays in IR8, Jhona and Basmati) mutation frequency is more than additive (synergistic) but these treatments are decisively less efficient because of their relatively high injurious effects in the M₁. generation. EMS induces more albinas than gamma-rays do. The mutational spectrum patterns induced by gamma-rays and DES are alike. In general, combination treatments tend to increase the frequency of albinas over other types of chlorophyll mutants.     

Short-term changes in carbon-isotope discrimination identify transitions between C3 and C4 carboxylation during Crassulacean acid metabolism

Short-term measurements of instantaneous carbon-isotope discrimination have been determined from mass-spectrometric analyses of CO₂ collected online during gas exchange for the epiphytic bromeliad Tillandsia utriculata L. Using this technique, the isotopic signature of CO₂ exchange for each phase of Crassulacean acid metabolism (CAM) has been characterised. During night-time fixation of CO₂ (Phase I), discrimination () ranged from 4.4 to 6.6, equivalent to an effective carbon-isotope ratio (¹³C) of –12.3 to –14.5 versus Pee Dee Belemnite (PDB). These values reflected the gross photosynthetic balance between net CO₂ uptake and refixation of respiratory CO₂, characteristic of CAM in the Bromeliaceae. When for the relative proportion of external (p _a ) and internal (p _i) CO₂ is taken into account, calculated p _i/p _a decreased during the later part of the dark period from 0.68 to 0.48. Measurements of during Phase II, early in the light period, showed the transition between C₄ and C₃ pathways, with carboxylation being increasingly dominated by ribulose bisphosphate carboxylase (Rubisco) as increased from 10.5 to 21.2 During decarboxylation in the light period (Phase III), CO₂ leaked out of the leaf and the inherent discrimination of Rubisco was expressed. The value of calculated from on-line measurements (64.4) showed that the CO₂ lost was considerably enriched in ¹³C, and this was confirmed by direct analysis of the CO₂ diffusing out into a CO₂-free atmosphere ( ¹³C = + 51.6 versus PDB). Instantaneous discrimination was characteristic of the C₃ pathway during Phase IV (late in the light period), but a reduction in showed an increasing contribution from phosphoenolpyruvate carboxylase. The results from this non-invasive technique confirm the observations that double carboxylation involving both phosphoenolpyruvate carboxylase and Rubisco occurs during the transient phases of CAM (II and IV) in the light period.Abbreviations and Symbols CAM Crassulacean acid metabolism - H⁺ (dawn-dusk) variation in titratable acidity - ¹³C carbonisotope ratio of plant organic material, relative to Pee Dee Belemnite (vs. PDB) - discrimination against ¹³CO₂, - p _i, p _a internal, external partial pressures of CO₂ - Rubisco ribulose1,5-bisphosphate carboxylase - PAR photosynthetically active radiation - PEPCase phosphoenolpyruvate carboxylase We are grateful for financial support in respect of research grants (GR3/5360, GR3/6419) and a studentship awarded by the Natural Environment Research Council, UK.     

Occurrence of pppApp-synthesizing activity in actinomycetes and isolation of purine nucleotide pyrophosphotransferase

The occurrence of adenosine 5-triphosphate-3-diphosphate-synthesizing activity was detected in five strains of actinomycetes; Streptomyces morookaensis, Streptomyces aspergilloides, Streptomyces hachijoensis, Actinomyces violascens and Streptoverticillium septatum, out of 825 strains of actinomycetes, bacteria, fungi and imperfecti. Purine nucleotide pyrophosphotransferase were extracellularly excreted associating with the cell growth, and were purified partially or to apparent homogeniety from the culture filtrate. The enzymes are a monomeric protein with a molecular weight of 18000–26000 and synthesize adenosine, guanosine and inosine 5-phosphate (mono, di or tri)-3-diphosphate such as pApp, ppApp, pppApp, pGpp, ppGpp, pppGpp and pppIpp by transferring a pyrophosphoryl group from the 5-position of ATP, dATP and pppApp to the 3-position of purine nucleotides in the presence of a divalent cation and in alkaline state.Abbreviations pppApp adenosine 5-triphosphate 3-diphosphate - ppApp adenosine 5-diphosphate 3-diphosphate - pApp adenosine 5-monophosphate 3-diphosphate - pppGpp guanosine 5-triphosphate 3-diphosphate     

Amino acid deletions in loop C of the chlorophyll a-binding protein CP47 alter the chloride requirement and/or prevent the assembly of photosystem II

The chlorophyll a-binding protein CP47 directs excitation energy to the reaction center of photosystem II (PSII) during oxygenic photosynthesis and has additional structural and functional roles associated with the PSII water-oxidizing complex. Oligonucleotide-directed mutagenesis was employed to study loop C of CP47 (approximately Trp-162 to Gly-197) which faces the thylakoid lumen. Five short amino acid deletion strains, (S169–P171), (Y172–G176), (G176–P180), (E184–A188) and (F190–N194), were created that span this domain. The deletion between Gly-176 and Pro-180, located around the middle of loop C, produced an obligate photoheterotroph that could not assemble functional PSII centers. The deletions in mutants (S169–P171) and (Y172–G176) reduced PSII levels to 20% of the control and thus impaired photoautotrophic growth. In contrast, mutants (E184-A188) and (F190–N194) were photoautotrophic even though the number of photosystems was decreased by 50%. All PSII complexes assembled in the deletion strains had an increased susceptibility to photoinactivation and deletion of Glu-184 to Ala-188 prevented photoautotrophic growth under chloride-limiting conditions. Furthermore, the removal of the extrinsic PSII-O, PSII-U and PSII-V proteins from mutants (E184–A188) and (F190–N194) reduced the rates of oxygen evolution and, in the strains lacking either the PSII-O or PSII-V proteins, also increased the photoautotrophic doubling times. These effects were greater in mutant (E184–A188) than in mutant (F190–N194) and the order of importance for the removal of the extrinsic proteins was found to be PSII-V PSII-O > PSII-U.     

Genetic analysis of two wheat cultivars, ‘Sonalika’ and ‘WL 711’ for reaction to leaf rust (Puccinia recondita)

Summary Genetic analysis for leaf rust reaction of two widely adapted cultivars, Sonalika and WL 711, has been done using 21 near isogenic Lr lines and rust culture IL004 — avirulent on the two cultivars and all the Lr lines used. The segregation pattern in the F₂ generation indicated the presence of a recessive gene in Sonalika and of a dominant gene in WL 711. These genes in cultivars Sonalika and WL 711 have been identified as Lr 11 and Lr 13, respectively. Gene Lr 13 is no longer effective in WL 711 but it continues to give field resistance in the backgrounds of Chris, Prelude and Thatcher. There has been no significant change in the virulence spectrum of the leaf rust pathogen in India with the release of WL 711. High susceptibility of WL 711 seems to be due to the evolution of more aggressive forms of the pathogen to this cultivar. The gene Lr 11, which behaves as a recessive in Sonalika, was effective against leaf rust when this cultivar was released. The high susceptibility of Sonalika is probably due to an increase in the frequency of race 77 virulent on Lr 11. Lr 11 has shown a dominance reversal in the background of Sonalika. Present results suggest that interaction of resistance genes with the background genotype must be studied for their effective use in breeding programme.     

The membrane potential in whole cells of Methanobacterium thermoautotrophicum

of whole cells of Methanobacterium thermoautotrophicum was estimated under varying conditions using an electrode sensitive to the lipophilic cation tetraphenylphosphonium chloride (TPP⁺). Since was found to be extremely sensitive to air, a special reaction vessel was developed to maintain strict anaerobiosis. The cells took up TPP⁺ under energization by H₂ and CO₂ thus allowing to calculate the from the distribution of TPP⁺ inside and outside the cells. The unspecific uptake of deenergized cells was around 10% of the total uptake of energized cells. TPP⁺ itself slightly diminished the , but had no effect on the formation of methane. Typical values of were in the range of-150 to-200 mV. showed a quantitative dependence on both the electron donor H₂ and the electron acceptor CO₂. NaCl stimulated the extent of the , whereas KCl slightly diminished it. Valinomycin resulted in a linear decline of , whereas the methane production rate was only slightly affected. In contrast, monensin reduced both methanogenesis and .Abbreviations pmf proton motive force - membrane potential - TPP⁺ tetraphenylphosphonium (chloride salt) - TPMP⁺ triphenylmethylphosphonium (chloride salt, if not otherwise indicated) - d.w. dry weight - t _d doubling time - PVC polyvinylchloride     

Hexahydroindanone derivatives of steroids formed by Rhodococcus equi

Summary A mutant strain of Rhodococcus equi accumulates three metabolites from the androst-4-ene-3,17-dione or from its degradation intermediate, 3a-H-4(3'-propionic acid)-7a-methylhexahydro-1,5-indanedione (MEPHIP). These three metabolites are: 3a-H-4a(3'-propionic acid)-5-hydroxy-7a-methylhexahydro-1-indanone--lactone (HIL); 3a-H-4(3'-trans acrylic acid)-5-hydroxy-7a-methylhexahydro-1-indanone (^2'-5-hydroxy-MEPHIP); and 3a-H-4(3'-hydroxy-3'-propionic acid)-5-hydroxy-7a-methylhexahydro-1-indanone (3'-hydroxy-HIL). The behaviour of this mutant allows us to propose a pathway for degradation of the intermediates, methylperhydroindanone propionates. However, during this degradation, the side-chain propionate was eliminated by a-oxidation mechanism. Offprint requests to: A. Miclo     

The electrochemical proton gradient and its influence on citrate uptake in tonoplast vesicles of Hevea brasiliensis

The relationship between the electrochemical proton gradient, _H+ ^– , and citrate transport has been studied in tonoplast vesicles from Hevea brasiliensis (the rubber tree). Vesicles were generated from lyophilized samples of fresh vacuoles obtained from the latex sap. Methylamine was used to measure intravesicular pH and lipophilic ions to determine the electrical potential difference () across the tonoplast. When incubated at pH 7.5 in the absence of ATP, the tonoplast vesicles showed a pH of 0.6 units (interior acid) and a of about-100 mV (interior negative). This potential is thought to be made up of contributions from an H⁺ diffusion potential, diffusion potentials from other cations and a Donnan potential arising from the presence of internal citrate. In the presence of 5 mol m^-3 MgATP the pH was increased to about 1.0 unit and the to about-10 mV. Under these conditions the proton-motive force ( _{p H+} ^– /F) became positive and reached +50 mV. These effects were specific to MgATP (ADP and Mg²⁺ having no significant effect) and were prevented by the protonophore p-trifluoromethoxycarbonylcyanidephenylhydrazone (FCCP). Citrate uptake by the vesicles was markedly stimulated by MgATP; ADP and Mg²⁺ again had no effect. Nigericin greatly increased pH and this was associated with a large increase in citrate accumulation. The results indicate that the vesicle membrane possesses a functional H⁺-translocating ATPase. The _H+ ^– generated by this ATPase can be used to drive citrate uptake into the vesicles. The properties of the tonoplast vesicles are compared with those of the fresh latex vacuoles.Abbreviations _H+ ^– electrochemical proton gradient - electrical potential difference across membrane - p proton-motive force ( _H+ ^– /F) - FCCP p-trifluoromethoxycarbonylcyanidephenylhydrazone - TPMP⁺ triphenylmethylphosphonium ion     

High-level Expression of Maize <Emphasis Type="Italic">γ</Emphasis>-zein Protein in Transgenic Soybean (<Emphasis Type="Italic">Glycine max</Emphasis>)

Primers were developed for 118 microsatellites isolated from grape (Vitis vinifera) genomic libraries enriched for (AC)n repeats. Only one microsatellite sequence matched other grape SSR-sequences in the GeneBank database. Genotyping was carried out in the parental lines and four offspring of two pseudo-test-cross populations, Cabernet Sauvignon x Seyval and Chardonnay x Bianca, and a further six other grape genotypes (V. vinifera Sultanina, Merlot, Syrah, Müller-Thurgau, Vitis Regent and V. riparia Gloire de Montpellier). A total of 108 microsatellites showed easily scorable alleles and 100 of them segregated according to a configuration suitable for mapping in either cross. A further 8 SSRs, although unsuitable for mapping in those crosses, showed polymorphism in the other genotypes tested. This set of markers was used, along with 75 microsatellites of other repeat-types, to fingerprint 46 offspring of the cross Chardonnay x Bianca. For each full-sib, individual heterozygosity and distance in repeat units between pairs of alleles at each locus (mean d²) were calculated as a tool for predicting highly outbred recombinant individuals. Six microsatellites with segregation ratios significantly distorted towards the lack of homozygous sibs were identified and mapped to linkage groups LG 3 and LG 5. Estimation of heterozygosity at genome-wide level and genotyping at loci for which homozygous sibs are discriminated against are discussed for marker-assisted background selection in outcrossing grapevines.     

Genetic differentiation in carbon isotope discrimination and gas exchange in Pseudotsuga menziesii

Patterns of genetic variation in gas-exchange physiology were analyzed in a 15-year-old Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) plantation that contains 25 populations grown from seed collected from across the natural distribution of the species. Seed was collected from 33°30 to 53°12 north latitude and from 170 m to 2930 m above sea level, and from the coastal and interior (Rocky Mountain) varieties of the species. Carbon isotope discrimination () ranged from 19.70() to 22.43() and was closely related to geographic location of the seed source. The coastal variety (20.50 (SE=0.21)) was not significantly different from the interior variety (20.91 (0.15)). Instead, most variation was found within the interior variety; populations from the southern Rockies had the highest discrimination (21.53 (0.20)) (lowest water-use efficiency). Carbon isotope discrimination (), stomatal conductance to water vapor (g), the ratio of intercellular to ambient CO₂ concentration (c_i/c_a), and intrinsic water-use efficiency (A/g) were all correlated with altitude of origin (r=0.76, 0.73, 0.74, and –0.63 respectively); all were statistically significant at the 0.01 level. The same variables were correlated with both height and diameter at age 15 (all at P0.0005). Observed patterns in the common garden did not conform to our expectation of higher WUE, measured by both A/g and , in trees from the drier habitats of the interior, nor did they agree with published in situ observations of decreasing g and with altitude. The genetic effect opposes the altitudinal one, leading to some degree of homeostasis in physiological characteri tics in situ.     

Reaction centers from three herbicide resistant mutants of Rhodobacter sphaeroides 2.4.1: Kinetics of electron transfer reactions

Electron transfer rates were measured in RCs from three herbicide-resistant mutants with known amino acid changes to elucidate the structural requirements for last electron transfer. The three herbicide resistant mutants were IM(L229) (Ile-L229 Met), SP(L223) (Ser-L223 Pro) and YG(L222) (Tyr-L222 Gly). The electron transfer rate D⁺Q_A ^-Q_BD⁺Q_AQ_B (k _AB) is slowed 3 fold in the IM(L229) and YG(L222) RCs (pH 8). The stabilization of D⁺Q_AQ_B ^- with respect to D⁺Q_AQ_B ^- (pH 8) was found to be eliminated in the IM(L229) mutant RCs (G⁰ 0 meV), was partially reduced in the SP(L223) mutant RCs (G⁰=–30 meV), and was unaltered in the YG(L222) mutant RCs (G⁰=–60 meV), compared to that observed in the native RCs (G⁰=–60 meV). The pH dependences of the charge recombination rate D⁺Q_AQ_B ^-DQ_AQ_B (k _BD) and the electron transfer from Q_A ^- (k _QA ^-Q_A) suggest that the mutations do not affect the protonation state of Glu-L212 nor the electrostatic interactions of Q_B and Q_B ^- with Glu-L212. The binding affinities of UQ₁₀ for the Q_B site were found in order of decreasing values to be native IM(L229) > YG(L222) SP(L223). The altered properties of the mutant RCs are used to deduce possible structural changes caused by the mutations and are dicscussed in terms of photosynthetic efficiency of the herbicide resistant strains.Abbreviations Bchl bacteriochlorophyll - Bphe bacteriopheophytin - cholate 3,7,12-trihydroxycholanic acid - D donor (bacteriochlorophyll dimer) - EDTA ethylenediamine tetraacetic acid - Fe²⁺ non-heme iron atom - LDAO lauryl dimethylamine oxide - PS II photosystem II - Q_A and Q_B primary and secondary quinone acceptors - RC bacterial reaction center - Tris tris(hydroxymethyl)aminomethane - UQ₀ 2,3-dimethoxy-5-methyl benzoquinone - UQ₁₀ ubiquinone 50     

13C discrimination during CO2 assimilation by the terrestrial biosphere

Estimates of the extent of the discrimination against¹³CO₂ during photosynthesis (_A) on a global basis were made using gridded data sets of temperature, precipitation, elevation, humidity and vegetation type. Stomatal responses to leaf-to-air vapour mole fraction difference (D, leaf-to-air vapour pressure difference divided by atmospheric pressure) were first determined by a literature review and by assuming that stomatal behaviour results in the optimisation of plant water use in relation to carbon gain. Using monthly time steps, modelled stomatal responses toD were used to calculate the ratio of stomatal cavity to ambient CO₂ mole fractions and then, in association with leaf internal conductances, to calculate _A. Weighted according to gross primary productivity (GPP, annual net CO₂ asimilation per unit ground area), estimated _A for C₃ biomes ranged from 12.9 for xerophytic woods and shrub to 19.6 for cool/cold deciduous forest, with an average value from C₃ plants of 17.8. This is slightly less than the commonly used values of 18–20. For C₄ plants the average modelled discrimination was 3.6, again slightly less than would be calculated from C₄ plant dry matter carbon isotopic composition (yielding around 5). From our model we estimate that, on a global basis, 21% of GPP is by C₄ plants and for the terrestrial biosphere as a whole we calculate an average isotope discrimination during photosynthesis of 14.8. There are large variations in _A across the globe, the largest of which are associated with the precence or absence of C₄ plants. Due to longitudinal variations in _A, there are problems in using latitudinally averaged terrestrial carbon isotope discriminations to calculate the ratio of net oceanic to net terrestrial carbon fluxes.     

Disulfide linked αoLH-gelonin conjugate failed to recombine with βoLH subunit to generate bioeffective hormonotoxin

Since, linking of ovine luteinizing hormone (oLH) to ribosome inactivating protein gelonin (in oLH-gelonin conjugate) occur via the alpha-subunit, oLH, an attempt has been made to develop a universal hormonotoxin for selective targeting to specific cells in the gonads. Four different molar ratios of oLH and N-succinimidyl-3-(2-pyridyldithio) propionate (SPDP) were used to activate the epsilon amino (-NH₂) groups of oLH. The oLH-SPDP derivatives recombine to native beta subunit of oLH (oLH) and the purified recombinants retained substantial receptor binding, steroidogenic activity and immunoreactivity to native oLH. The disulfide linked oLH-S-S-gelonin conjugates prepared by SPDP method were purified by gel filtration chromatography and analysed by reverse-phase high performance liquid chromatography (RP-HPLC). In order to obtain specificity and bioeffectivity, the oLH-S-S-gelonin conjugates were allowed to recombine to native oLH and the recombination mixture was further purified by gel-filtration chromatography. The RP-HPLC analysis of these recombinants indicated that oLH-S-S-gelonin did not recombine to oLH. The failure of recombination may be due to the reasons. (i) The site of -NH₂ activation by SPDP may be different in the oLH than the native oLH. (ii) The activation site may be in close proximity to the annealing site which facilitates the recombination of -subunit but failured to reassociate to oLH-S-S-gelonin conjugate. (iii) The introduction of gelonin (30 kDa basic protein) might have induced some steric hinderence for oLH to recombine to the oLH site which might have been masked in oLH-S-S-gelonin conjugates. (Mol Cell Biochem120: 95–102, 1993)Abbreviations oLH ovine Luteinizing Hormone - oLH alpha subunit of oLH - oLH beta subunit of oLH - BSA Bovine Serum Albumin - DTT Dithiothreitol - RP-HPLC Reverse Phase High Performance Liquid Chromatography - TSH Thyroid Stimulating Hormone - FSH Follicle Stimulating Hormone - LH Luteinizing Hormone - eCG equine Chorionic Gonadotropin - DMEM Dulbecco's Modified Eagles Medium - HEPES 4-(2-Hydroxyethyl)-1 Piperazine Ethane Sulfonic acid - PAP Pokeweed Antiviral Protein - RIA Radioimmunoassay - hCG human Chorionic Gonadotropin - TRH Thyrotropin Releasing Hormone - CRF Corticotropin Releasing Factor - hPL human Placental Lactogen - TFA Trifluroacetic Acid - oLH-SPDP SPDP activated derivative of oLH     

Electrochemical potential of protons in vesicles reconstituted from purified,proton-translocating adenosine triphosphatase

Summary Measurements were made of the difference in the electrochemical potential of protons ( ) across the membrane of vesicles reconstituted from the ATPase complex (TF ₀ ·F ₁) purified from a thermophilic bacterium and P-lipids. Two fluorescent dyes, anilinonaphthalene sulfonate (ANS) and 9-aminoacridine (9AA) were used as probes for measuring the membrane potential () and pH difference across the membrane ( pH), respectively.In the presence of Tris buffer the maximal and no pH were produced, while in the presence of the permeant anion NO ₃ ^– the maximal pH and a low were produced by the addition of ATP. When the ATP concentration was 0.24mm, the was 140–150 mV (positive inside) in Tris buffer, and the pH was 2.9–3.5 units (acidic inside) in the presence of NO ₃ ^– . Addition of a saturating amount of ATP produced somewhat larger and pH values, and the attained was about 310 mV.By trapping pH indicators in the vesicles during their reconstitution it was found that the pH inside the vesicles was pH 4–5 during ATP hydrolysis.The effects of energy transfer inhibitors, uncouplers, ionophores, and permeant anions on these vesicles were studied.