A procedure for the isolation of malic enzyme from muscle tissue of the roundworm Ascaris suum is described. The fractionation method yields relatively large quantities of the enzyme,with a specific activity of fifteen moles of malate converted to pyruvate and carbon dioxide per min per mg at 25º. Homogeneity was established with analytical ultracentrifugation, zone electrophoresis, isoelectric focusing, and rechromatography. The molecular weight of the enzyme was 250,000, and it is dissociated under several conditions into four identical monomers of 64,000 daltons. The enzyme exists as a single electrophoretic form and prefers manganous and NAD over other cations and NADP. Ammonium sulfate competes with manganous for the active site and titration with DTNB yields eight thiol groups per mole. Titration of the first four thiol groups is accompanied by a complete loss in enzyme activity. Equilibrium dialysis, product inhibition, and initial velocity studies suggest a rapid-equilibrium random sequential mechanism for the Ascaris suum malic enzyme. The presence of 1.3 binding sites per subunits was determined for L-ma late. Antisera prepared against A. suum malic enzyme reacted to a small extent with the NAD malic enzymes from two free-living nematodes, Panarellus redivivus and Turbatrix aceti. A correlation coefficient of 0.911 was obtained upon comparing the amino acid composition of A. suum and E. coli malic enzymes. Some sequence homology is predicted between these malic enzymes. The physiological interpretation favors the binding of malate initially, with the subsequent addition of NAD to the enzyme.

Proefschrift - Utrecht.

Thesis (Ph.D.) -- McMaster University, 2001. / Includes bibliographical references. Also available via World Wide Web.

Thesis (Ph.D.)--University of Wollongong, 2008. / Typescript. Includes bibliographical references.

The diploma thesis is focused on the study of the influence of cultivation conditions and arrangement for the production of extracellular polymeric substances by using yeast-like microorganism Aureobasidium pullulans. In the theoretical part a brief description of A. pullulans, its use in biotechnology and produced exobiopolymers, especially pullulan and poly-L-malic acid are presented. The first aim of the experimental part was to set the most appropriate cultivation conditions for A. pullulans CCM 8182. Growth and production properties in optimum conditions were compared with cultivation on waste substrates - oat bran, buckwheat husks, apple fiber and others. Waste substrates can be used as cheap nutrient sources which enable reducing cost of potential biotechnological production. As a further part of this work, optimization of HPLC/RI method for analysis of exobiopolymers has been done. Optimal mobile phase composition and chromatography conditions were proposed. Column Roa organic acid H+ was the most suitable for simultaneous separartion of glucose and malic acid. Before HPLC analysis hydrolysis of polymers was done. Sulphuric acid (5 mmol/L) was used as a mobile phase at flow rate 0.5 mL/min and temperature 60 °C. The highest production of pullulan occurred using oat bran as a substarate (13.03 g/L) at an initial pH 7.5. Maximum production of poly-L-malic acid was observed during the cultivation on apple peels (2.89 g/L) at pH 6. It was found that the higher production of poly-L-malic acid occurred at pH 6, while higher production of pullulan was at pH 7.5.

Thirty-two lactating cows were assigned at random to four treatments of malic acid to determine if these levels had an effect on milk production, milk composition, feed intake, and efficiency of feed utilization. Malic acid allotment for each treatment consisted of 1) 15.4, 2) 11.6, 3) 7.7, and 4) 0 grams of malic acid fed per kilogram of concentrate. Concentrate was fed according to production at a rate of one kilogram per two kilograms of milk in excess of 9.1 kilograms of milk per cow daily. Alfalfa hay was fed free choice and corn silage at a rate of 11.4 kilograms daily. The cows were on the trial for 8 weeks. Intake of concentrates, silage, dry matter, and digestible energy was highest for cows receiving the 11.6 g level of malic acid. These intakes were significantly higher than for the 7.7 g level but not for the other treatments. However, cows on the 7.7 g level consumed only slightly less feed than control cows. There was no significant effect on hay or crude protein intake. Production of total milk, fat corrected milk, and milk fat was significantly higher for cows receiving the 11.6 g level of malic acid than from the 7.7 g level or control cows. Production of protein solids-not-fat was significantly higher for the 11.6 g level than from the 7.7 g level and approached this level of significance when the 11.6 g level was compared to the controls. Cows receiving the 11.6 g level of malic acid were significantly more efficient in converting dry matter or digestible energy from feed into milk than were the controls. Intakes and production of cows on the 15.4 g level was slightly less than for the 11.6 g level.

京都大学 / 新制・論文博士 / 博士(農学) / 乙第13458号 / 論農博第2899号 / 新制||農||1088(附属図書館) / 学位論文||R3||N5362(農学部図書室) / (主査)教授 小川 順, 教授 阪井 康能, 教授 栗原 達夫 / 学位規則第4条第2項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

No / This review summarises the recent developments in the synthesis and applications of polymers derived from malic acid. There has been an increased interest in the design of sustainable and biodegradable polymers as a result of the drive to use renewable feedstocks as an alternative to petrochemicals in addition to their significant potential in biomedical applications. Synthetic strategies to access polymers from malic acid based on both condensation and ring-opening polymerization, across a broad range of conditions, are reviewed along with their advantages and limits. The role that such materials are studied for in biomedical applications is discussed, and their environmental impact based on the biodegradability of the malic polymer backbone is outlined. / The Royal Society, EPSRC, BBSRC

Delivery of biologically relevant ions such as drugs, neurotransmitters and hormones have been recognized as powerful a tool to control physiology of animals and plants for research purposes and practical applications. In the plant research community, ions are most commonly delivered as part of a solvent by soaking, spraying, pipetting or by adding to the soil. These methods have low control of the delivery dynamics and quantity of ion uptake. These issues motivated the development of the Organic Electronic Ion Pump (OEIP), which delivers only ions of interest by applying an external electric field through a polyelectrolyte membrane of high fixed charge concentration. A miniaturized, implantable version of the OEIP based on capillary fibres (c-OEIP), where the polyelectrolyte is enclosed in a capillary, enabled even higher precision of the delivery. In this master thesis, c-OEIP has been applied in the tropical plant Kalanchoe Blossfeldiana, chosen due to its characteristic skill to gradually learn to save water: while maturing it shifts to night time photosynthesis and transpiration, called Constitutive Crassulacean Acid Metabolism. A better understanding of this metabolism and water saving ability could guide engineering of enhanced drought tolerance in crop plants, which is motivated by the increasing global warming. One of the biologically relevant ions that is potentially involved in this water-saving learning process is the malate ions. The aim of this thesis is to test the hypothesis that c-OEIP is able to deliver malate ions to cause a reduction in stomatal conductance and transpiration of intact leaves of Kalanchoe Blossfeldiana. To test this hypothesis, firstly, the capillary-based OEIP were fabricated using polyimide coated glass capillaries filled with AETMAC polyelectrolyte. The ability of these devices to deliver malic acid (MA) was verified by using current-voltage characterisation during loading and delivery of MA. Secondly, the setup for MA delivery with c-OEIP to intact kalanchoe leaf was developed, optimising the insertion method to minimize the wounding of the plant and increase assay reproducibility. Finally, the MA was delivered to intact kalanchoe leaves via c-OEIP, where the plant transpiration response was evaluated using standard gas exchange porometer and also novel infrared camera, as plant temperature can be correlated with plant transpiration status. The results indicate that c-OEIP can deliver MA and trigger reduction of transpiration of young kalanchoe leaves, supporting the hypothesis that malate ions act to reduce stomatal conductance, potentially conveying a feedback message from the mesophyll to the guard cells. / <p>Examensarbetet är utfört vid Institutionen för teknik och naturvetenskap (ITN) vid Tekniska fakulteten, Linköpings universitet</p>

Stomata are small pores that are located on the surface of epidermal leaves, and they can regulate the uptake of CO2 and prevent water lose by opening and closing the pores. Stomata of plants can be regulated by external condition such as CO2, biotic and abiotic stresses and internal factors. CAM (crassulacean acid metabolism) plants adapt to hot and dry environments by closing stomata during the day and opening stomata during the cool night. However, it is still unclear how CAM plants open their stomata during the night and close them during the day. In this study, a number of factors were evaluated for their potential roles in promoting stomatal opening in the model CAM plant Kalanchoe laxiflora. Citrate is an important organic acid and it accumulates during the night in CAM plants. It is shown in this study that citrate promoted stomatal opening in detached leaf epidermis of Kalanchoe laxiflora. Further, the cytokinin zeatin is also shown to stimulate stomatal opening in detached leave of Kalanchoe laxiflora. Melatonin is an important regulator of circadian rhythms in mammals and has been implicated in regulation of plant abiotic stress responses. Melatonin was detected in the leaves of Kalanchoe laxiflora. It promoted stomatal opening in detached epidermis of Kalanchoe laxiflora. Together, these results suggest that stomata of Kalanchoe laxiflora respond to citrate and malate which are the main organic acids accumulate during nighttime and also to some signaling molecules (zeatin, melatonin, and serotonin) by opening stomata during dark period.