Photoinactivation of δ-aminolevulinic acid dehydratase from maize by flavin mononuclotide

The -aminolevulinic acid dehydratase activity was irreversibly inactivated by irradiation of the enzyme in presence of flavin mononucleotide. The loss of enzyme activity was dependent on time of irradiation, concentration of FMN and intensity of irradiance. It required oxygen and was markedly enhanced in heavy water. The presence of levulinic acid (a competitive inhibitor of -ALAD) during irradiation prevented the inactivation considerably indicating photooxidative damage at or near the active site. Superoxide dismutase, sodium benzoate and sodium formate offered no protection, but singlet oxygen quenchers like azide and tryptophan were effective. NADH, electron donor to excited flavins, also prevented the loss of enzyme activity. These results indicate that singlet oxygen produced by light absorption of FMN was responsible for the photooxidative inhibition of the enzyme.Abbreviations ALAD -aminolevulinic acid dehydratase - FMN flavin mononucleotide - O₂ ^- superoxide - H₂O₂ hydrogen peroxide - 10₂ singlet oxygen - LA levulinic acid - PBG porphobilinogen - BSA bovine serum albumin - BME 2-mercaptoethanol - SOD superoxide dismutase - pHMB para-hydroxymercuribenzoate - DTT dithiothreitol - FAD flavin adenine dinucleotide - NADH nicotinamide adenine dinucleotide     

Influence of zinc-saccharide complexes on some haematological parameters in rats

The effects of three recently synthesized zinc-saccharide complexes (zinc-fructose, zinc-galactose and zinc-glucose) on blood -aminolevulinic acid dehydratase (ALAD) activity, glutathione (GSH), zinc-protoporphyrin (ZPP) and urinary -aminolevulinic acid (ALA) levels have been investigated to ascertain the utility of these complexes as zinc supplements and as preventive agents against lead intoxication in rats.     

Capacity for chlorophyll synthesis in heat-bleached 70S ribosome-deficient rye leaves

The leaves of young rye plants (Secale cereale L.) grown at 32° were deficient in chlorophyll and in chloroplastic rRNA as compared to those grown at 22°, which developed normally. Both chlorophyll accumulation and the formation of plastidic rRNA were largely restored at 32° when the plants were transfered several times for 1 h per day to 22°. In the chlorotic 32°-grown rye leaves the in vivo activity of -aminolevulinate synthetase was very low. Aminolevulinate dehydratase however, exhibited high activity in extracts from 32°-grown leaves and was localized in the plastid fraction isolated from the chlorotic leaf tissue. After application of -aminolevulinic acid to chlorotic parts of leaves growing at 32°, protochlorophyll(ide) was formed and accumulated in the dark. In the light, the protochlorophyll(ide) was photooxidized at 32°. The results suggest a cytoplasmic site of synthesis for the series of enzymes converting -aminolevulinate to protochlorophyll(ide). It is concluded that an inhibition of -aminolevulinate synthetase and the photooxidation of protochlorophyll(ide) or chlorophyll are responsible for the chlorosis of the leaves at 32°.Abbreviations ALA -aminolevulinic acid - ALAD -aminolevulinate dehydratase - ALAS -aminolevulinate synthetase     

Heme metabolism in promastigotes of Leishmania donovani

Promastigotes of Leishmania donovani (Dd-8 strain) showed presence of important key enzymes of heme synthesizing (d-aminolevulinic acid synthase and ferrochelatase) and degrading (heme oxygenase and biliverdin reductase) systems, classical leishmanicidal drugs viz allopurinol, amphotericin B, pentamidine and CDRI compound 93/202 inhibited the heme oxygenase activity of the parasite, whereas, -aminolevulinic acid synthase activity practically remained unaffected. The Km, Vmax ad pH values of heme oxygenase of promastigotes were found to be 1666 M hemin, 625 nmol of bilirubin formed h-1 mg protein-1 and 7.5 respectively. The findings suggest the presence and importance of heme metabolism in the de novo synthesis of different hemoproteins of the Leishmania parasite as well as the detoxification and its defence against biological insults.     

The role of vanadium in green plants

In a series of experiments, it is demonstrated that the trace element vanadium (4·10^-7 g-at/l as NH₄VO₃) has a considerable positive influence on the synthesis of -aminolevulinic acid (-ALA) in the autotrophically growing green algaChlorella pyrenoidosa, the effect being visible by an enhanced output of the amino acid into the culture medium in presence of levulinic acid (LA). The level of intracellularly accumulated -ALA, however, is not changed in presence of the metal. The V-effect on exogenous found -ALA is suppressed, when LA is added to the nutrient medium at low pH (pH 5), although V-uptake into the algal cells is not disturbed by LA. As demonstrated in culture media with various nitrogen sources (urea, partially hydrolized urea, ammonium salts), the development of the pH during the cultivation time is important for the presentation of the V-effect on -ALA. It is suggested that vanadium acts as a catalyst in the conversion of 4,5-dioxovaleric acid to -ALA by transamination.Abbreviations -ALA -aminolevulinic acid - LA levulinic acid - DOVA 4,5-dioxovaleric acid     

Therapeutic potential of meso 2,3-dimercaptosuccinic acid or 2,3-dimercaptopropane 1-sulfonate in chronic arsenic intoxication in rats

The therapeutic efficacy of two thiol chelators, meso 2,3-dimercaptosuccinic acid (DMSA) or 2,3-dimercaptopropane sulfonate (DMPS) in treating chronic arsenic intoxication was investigated in male rats. Both the chelators were effective in promoting urinary arsenic excretion and restoring arsenic induced inhibition of blood -aminolevulinic acid dehydratase activity and hepatic glutathione level. Elevation of urinary -aminolevulinic acid excretion and arsenic concentration in blood, liver and kidneys were reduced significantly by both the chelators. Histopathological lesions induced by arsenic were also effectively reduced by the above chelators. DMSA being more effective than DMPS. The results suggest DMSA and DMPS to be effective antidotes for treating chronic arsenic toxicity in experimental animals.     

δ-Toxin,unlike melittin,has only hemolytic activity and no antimicrobial activity: Rationalization of this specific biological activity

The antimicrobial activity of a synthetic peptide corresponding to -hemolysin had been examined. The peptide didnot exhibit antimicrobial activity against gram negative and gram positive micro-organisms unlike other hemolytic peptides like melittin. This lack of antibacterial activity arises due to the inability of -hemolysin to perturb the negatively charged bacterial cell surface and permeabilize the bacterial plasma membrane. However, the red blood cell surface has a structure considerably different from bacteria, and does not act as a barrier to molecules reaching the lipid membrane. Hence -toxin can lyse erythrocytes. Thus, the specificity in biological activity has been rationalized in terms of differences, in the interaction of the toxin with the bacterial and red blood cell surfaces.     

Organic nitrogen metabolism of phototrophic bacteria

Recent reviews dealing with phototrophic bacteria are concerned with bioenergetics, nitrogen fixation and hydrogen metabolism, synthesis of the photosynthetic apparatus and phylogeny/taxonomy. The organic N-metabolism of these phylogenetically diverse bacteria has last been reviewed in 1978. However, amino acid utilization and biosynthesis, ammonia assimilation, purine and pyrimidine metabolism and biosynthesis of -aminolevulinic acid as precursor of bacteriochlorophylls and hemes are topics of vital importance. This review focusses on utilization of amino acids as N- and C/N-sources, the pathways of purine and pyrimidine degradation, novel aspects of amino acid biosynthesis (with emphasis on branched-chain amino acids and -aminole-vulinic acid) and some aspects of ammonia assimilation and glutamate synthesis by purple bacteria, green sulfur bacteria and Chloroflexus aurantiacus.Abbreviations R Rhodospirillum - Rhb Rhodobacter - Rc Rhodocyclus - Rp Rhodopila - Rps Rhodopseudomonas     

Effect of organic and inorganic fertigation on yields, δ15N values, and δ13C values of tomato (Lycopersicon esculentum Mill. cv. Saturn)

We examined the effects of fertilizer application, especially the effects of fertigation and types of fertilizer (inorganic and organic) on yields and ¹⁵N and ¹³C values of tomato (Lycopersicon esculentum Mill. cv. Saturn). Fertigation is a method in which an appropriate diluted liquid fertilizer is applied to the plants each time they are drip-irrigated. We developed a method of organic fertigation using corn steep liquor (CSL) as the liquid fertilizer, because it is an industrial byproduct of cornstarch manufacture and can be used very effectively. We compared fruit yield, mineral content, ¹⁵N value, and ¹³C value of tomatoes grown under three different fertilizer treatments, basal dressing: basal dressing with granular chemical fertilizer; inorganic fertigation: fertigation with liquid chemical fertilizer; and organic fertigation: fertigaion with CSL. Mineral contents of tomatoes grown with basal dressing were generally lower than those grown under either fertigation treatment. These results indicated that yields and mineral contents were influenced more by the method of fertilizer application than by whether the fertilizers were inorganic or organic. There were, however, significant differences in the ¹⁵N values of tomato fruits grown under different types of fertilizer applications, especially between inorganic and organic fertilizers. The ¹⁵N value of the chemical fertilizer used for basal dressing was 0.81 ± 0.45{}, that of the chemical fertilizer for fertigation was 0.00 ± 0.04{}, and that of CSL was 8.50 ± 0.71{}. The ¹⁵N values of the soils reflected the ¹⁵N values of the fertilizers. Moreover, the ¹⁵N values of the fruits corresponded to the ¹⁵N values of the applied fertilizers. The ¹⁵N values were 3.18 ± 1.34{} in the fruits grown with a basal dressing of chemical fertilizer, 0.30 ± 0.61 in those grown under inorganic fertigation, and 7.09 ± 0.68 in those grown under organic fertigation. On the other hand, although the ¹³C values in the soil also reflected the ¹³C values of the applied fertilizers, there was no significant difference in the ¹³C values of fruits among the different treatments. In conclusion, because the ¹⁵N values of fertilizers correlated well with those of the fruits, it may be possible to use ¹⁵N values as an indicator of organic products.     

Overexpression of ornithine-δ-aminotransferase increases proline biosynthesis and confers osmotolerance in transgenic plants

Accumulationof proline is a way to increase tolerance to water stress in plants. Therefore,considerable attention has been devoted to optimise proline biosynthesis intransgenic plants. Glutamate and ornithine are both precursors of proline butwhile genes of the glutamate pathway were overexpressed in transgenic plants,no gene encoding an enzyme of the ornithine pathway was considered until now. Thepresent study aims to establish if the overexpression ofornithine--aminotransferase (-OAT) represents an additional wayto increase proline content. To achieve this goal, anArabidopsis -OAT cDNA was fused to the CaMV35Spromoter and introduced via Agrobacterium transformationinto Nicotiana plumbaginifolia. Overexpression of the-OAT cDNA in the analysed transgenic lines was linked to an increase in-OAT enzyme activity. The transgenic lines presenting high enzymaticactivity synthesized more proline than the control plants and showed a higherbiomass and a higher germination rate under osmotic stress conditions. Thesestudies reveal a new and efficient way to increase proline content in plantsand to enhance crop tolerance.     

Stimulation of crystallin synthesis in embryonic brain cells in culture

Summary Expression of -crystallin, a lens-specific protein, in 6-day-old chick embryonic brain cells was examined in situ and in vitro. The presence of minute amounts of -crystallin and its mRNA (-mRNA) in brain cells in situ was demonstrated by immunoblot and Northern blot analysis. In spreading cultures of the brain cells, -crystallin and -mRNA showed a significant increase from their in situ level. Immunohistological staining (peroxidase antiperoxidase) with monospecific anti-serum against -crystallin revealed that -producers were both epithelial cells and dendritic cells. Neither lentoidogenesis nor -crystallin expression was observed. Stimulation of -crystallin synthesis in cultured brain cells differed when compared with transdifferentiating cultures of neural retina cells. In the latter, -crystallin synthesis occurred concomitantly with differentiation of morphologically distinct lens cells containing -crystallin.     

Insulin secretion at high altitude in man

The effect of hypoxia on circulatory levels of insulin, its response to oral glucose administration (100 g) and changes in circadian rhythms of glucose as well as insulin were evaluated in euglycemic males at sea level (SL, 220 m) during their stay at high altitude (3500 m, SJ) and in high altitude natives (HAN).Basal glucose levels were not altered at high altitude but the rise in glucose ( glucose) after glucose load was significantly higher in SJ and HAN (p<0.01) as compared to SL values. An increase (p<0.01) both in basal as well as glucose induced rise in insulin secretion ( insulin) was observed at HA. The rise in insulin in SJ was significantly higher (p<0.01) than in HAN. This elevation in glucose and insulin levels was also evident at different times of the day. The circadian rhythmicity of glucose as well as insulin was altered by the altitude stress. The findings of the study show a rise in insulin level at HA but the hyperglycemia in the face of hyper-insulinism require the presumption of a simultaneous and dispropotionate rise of insulin antagonistic hormones upsetting the effect of insulin on glucose metabolism.Presented at International Conference of Biometeorology held at New Delhi from December 26–30, 1983.     

Effect of cycloheximide on the induction of tryptophan oxygenase mRNA by hydrocortisone invivo

Hydrocortisone increases rat liver tryptophan oxygenase mRNA activity as measured by a translational assay. Pretreatment of rats with cycloheximide thirty minutes before hydrocortisone administration largely prevents the hormonal induction of tryptophan oxygenase mRNA. Tryptophan oxygenase mRNA activity begins to increase after a lag of at least 30 to 60 minutes after hydrocortisone injection. These results suggest that the synthesis of intermediary protein(s) is required for the induction of tryptophan oxygenase mRNA by glucocorticoids.     

Central nervous versus total body thermosensitivity of the duck

Ducks were chronically implanted with thermodes in the POAH region, the lower brainstem or the vertebral canal. At thermoneutral conditions, lowering the temperature of the spinal cord (T_vc) or the lower brainstem (T_mb) stimulated metabolic heat production (M) with a subsequent rise of core temperature (T_c). Lowering the temperature of the POAH region (T_hy) induced a fall of T_c due to paradoxical activation of heat defence and, thus, induced slight to moderate general hypothermia depending on the cooling intensity. When T_hy was normalized, the hypothermia temporarily stimulated metabolic heat production until T_c was normalized. Cold sensitivity of the entire body, as revealed by the metabolic response to the hypothermia induced by preceding POAH cooling, and cold sensitivity of the spinal cord and the lower brainstem, as revealed by the metabolic response to local cooling, were quantified by calculating the quotient M/T from the maximum metabolic response and the experimentally induced drop of T_c, T_mb and T_vc. With lower brainstem cooling M/T_mbdid not exceed –0.4 W/(kg · C). With spinal cord cooling, M/T_vc did not exceed –0.6 W/(kg · C). The mean value of M/T_c after hypothermia induced by POAH cooling was –4.02 W/(kg · C). The results indicate that the cold sensitivity residing in the CNS of ducks represents only a small fraction of the entire cold sensitivity of the body.Presented at the Eighth International Congress of Biometeorology, 9–14 September 1979, Shefayim, Israel.     

Hydrogen isotope discrimination in higher plants: Correlations with photosynthetic pathway and environment

Summary The ratio of deuterium to hydrogen (expressed as D) in hydrogen released as water during the combustion of dried plant material was examined. The D value (metabolic hydrogen) determined on plant materials grown under controlled conditions is correlated with pathways of photosynthetic carbon metabolism. C₃ plants show mean D values of-132 for shoots and -117 for roots; C₄ plants show mean D values of -91 for shoots and-77 for roots and CAM plants a D value of-75 for roots and shoots. The difference between the D value of shoot material from C₃ and C₄ plants was confirmed in species growing under a range of glasshouse conditions. This difference in D value between C₃ and C₄ species does not appear to be due to differences in the D value (tissue water) in the plants as a result of physical fractionation of hydrogen isotopes during transpiration. In C₃ and C₄ plants the hydrogen isotope discrimination is in the same direction as the carbon isotope discrimination and factors contributing to the difference in D values are discussed. In CAM plants grown in the laboratory or collected from the field D values range from-75 to +50 and are correlated with ¹³C values. When deprived of water, the D value (metabolic hydrogen) in both soluble and insoluble material in leaves of Kalanchoe daigremontiana Hamet et Perr., becomes less negative. These changes may reflect the deuterium enrichment of tissue water during transpiration, or in field conditions, may reflect the different D value of available water in areas of increasing aridity. Whatever the origin of the variable D value in CAM plants, this parameter may be a useful index of the water relations of these plants under natural conditions.     

The effects of bromocriptine and prolactin on porphyrin biosynthesis and morphology in the female hamster Harderian gland

Porphyrin biosynthesis was examined in the Harderian gland of the female golden hamster by fluorometric assays of gland porphyrin content and by measuring the activity of a rate-limiting enzyme for haem biosynthesis, -aminolaevulinic acid synthase. Both porphyrin content and enzyme activity are high in normal female glands. Enzyme activity was lowered in females ovariectomised for 6 weeks, and both enzyme activity and porphyrin content were greatly lowered in ovariectomised females given the dopamine agonist bromocriptine; this suppression could be prevented by simultaneous prolactin administration. Bromocriptine (but not ovariectomy alone) also masculinised the morphology of the Harderian gland, resulting in the appearance of type II cells and polytubular complexes; again, the simultaneous administration of prolactin prevented masculinisation. The results support the hypothesis that while androgens have an inhibitory effect on porphyrin synthesis within this model, prolactin may have a major facilitatory role.Abbreviations ALA -aminolaevulinic acid - ALA-s -aminolaevulinate synthase     

Shoot δ15N correlates with genotype and salt stress in barley

Given a uniform N source, the ¹⁵N of barley shoots provided a genotypic range within treatments and a separation between control and salt-stress treatments as great as did ¹³C^*. Plant ¹⁵N has been represented in the literature as a bioassay of external source ¹⁵N and used to infer soil N sources, thus precluding consideration of the plant as a major cause in determining its own 8¹⁵N. We believe this to be the first report of plant ¹⁵N as a genetic trait. No mechanistic model is needed for use of ¹⁵N as a trait in controlled studies; however, a qualitative model is suggested for further testing.Symbol ¹⁵N (or ¹³C) the difference between: (1) the ratio of heavy to light isotopes of the element in a sample and (2) that of its reference standard     

Distribution of δ-aminolevulinic acid biosynthetic pathways among phototrophic bacterial groups

Two biosynthetic pathways are known for the universal tetrapyrrole precursor, -aminolevulinic acid (ALA). In the ALA synthase pathway which was first described in animal and some bacterial cells, the pyridoxal phosphate-dependent enzyme ALA synthase catalyzes condensation of glycine and succinyl-CoA to form ALA with the loss of C-1 of glycine as CO₂. In the five-carbon pathway which was first described in plant and algal cells, the carbon skeleton of glutamate is converted intact to ALA in a proposed reaction sequence that requires three enzymes, tRNA^Glu, ATP, Mg²⁺, NADPH, and pyridoxal phosphate. We have examined the distribution of the two ALA biosynthetic pathways among various genera, using cell-free extracts obtained from representative organisms. Evidence for the operation of the five-carbon pathway was obtained by the measurement of RNase-sensitive label incorporation from glutamate into ALA, using 3,4-[³H]glutamate or 1-[¹⁴C]glutamate as substrate. ALA synthase activity was indicated by RNase-insensitive incorporation of label from 2-[¹⁴C]glycine into ALA. The distribution of the two pathways among the bacteria tested was in general agreement with their previously established phylogenetic relationships and clearly indicates that the five-carbon pathway is the more ancient process, whereas the pathway utilizing ALA synthase probably evolved much later. The five-carbon pathway is apparently the more widely utilized one among bacteria, while the ALA synthase pathway seems to be limited to the subgroup of purple bacteria.Abbreviations ALA -aminolevulinic acid - DTT dithiothreitol - PALP pyridoxal phosphate - SDS sodium dodecyl sulfate - tricine N-tris-(hydroxymethyl)methylglycine     

Carbon isotope composition in relation to leaf gas exchange and environmental conditions in Hawaiian Metrosideros polymorpha populations

Summary Carbon isotope composition, photosynthetic gas exchange, and nitrogen content were measured in leaves of three varieties of Metrosideros polymorpha growing in sites presenting a variety of precipitation, temperature and edaphic regimes. The eight populations studied could be divided into two groups on the basis of their mean foliar ¹³C values, one group consisting of three populations with mean ¹³C values ca.-26 and another group with ¹³C values ca.-28. Less negative ¹³C values appeared to be associated with reduced physiological availability of soil moisture resulting from hypoxic conditions at a poorly drained high elevation bog site and from low precipitation at a welldrained, low elevation leeward site. Gas exchange measurements indicated that foliar ¹³C and intrinsic wateruse efficiency were positively correlated. Maximum photosynthetic rates were nearly constant while maximum stomatal conductance varied substantially in individuals with foliar ¹³C ranging from-29 to-24. In contrast with the patterns of ¹³C observed, leaf nitrogen content appeared to be genetically determined and independent of site characteristics. Photosynthetic nitrogenuse efficiency was nearly constant over the range of ¹³C observed, suggesting that a compromise between intrinsic water- and N-use efficiency did not occur. In one population variations in foliar ¹³C and gas exchange with leaf cohort age, caused the ratio of intercellular to atmospheric partial pressure of CO₂ predicted from gas exchange and that calculated from ¹³C to be in close agreement only in the two youngest cohorts of fully expanded leaves. The results indicated that with suitable precautions concerning measurement protocol, foliar ¹³C and gas exchange measurements were reliable indicators of potential resource use efficiency by M. polymorpha along environmental gradients.