Limonoate dehydrogenase

Enzymes in Bacteria. Limonoate dehydrogenases, which catalyze the conversion of XIV to XV, have been isolated from three species of bacteria by Hasegawa et al. (17,18,19). Each dehydrogenase has different characteristics (Table V). 

Activities of limonoate dehydrogenase (22), epoxidase (6) and deoxylimonate A-ring lactone hydrolase (6) have been demonstrated in Citrus, but they have not been isolated yet. 

Control of Juice Bitterness. A number of advances have been reported in this field since it was last reviewed (3). A commercial application of the cellulose acetate adsorption technique for the removal of limonin from citrus juices was undertaken (49). New sorbent gel forms of cellulose esters for adsorption of limonin were developed (50). Knowledge was gained that limonoids are biosynthesized in citrus leaves and translocated to the fruit (12) and that specific bioregulators can inhibit accumulation of XIV in citrus leaves (15). Additional studies were carried out on the use of neodiosmin to suppress limonin and other types of bitterness (30,51). The influence of extractor and finisher pressures on the level of limonin and naringin in grapefruit juice was reported (34). Also, further studies were conducted on the microbial sources and properties of limonoate dehydrogenase (52), the enzyme that converts XIV to XV and can be used to prevent limonin from forming in freshly expressed citrus juices (53). 

Hasegawa et al. (56) dj ected the enzymic conversion of 19-deoxylimonoic acid 3-methyl C ester to the 17-dehydro derivative by albedo tissue slices of Navel oranges. They isolated the product and identified it by TLC as the reaction product formed when the substrate was dehydrogenated by limonoate dehydrogenase (EC 1.1.1)... 

Hasegawa, S. Maier, V. P. Bennett, R. D. Detection of limonoate dehydrogenase activity in albedo tissues of Citrus sinensis. Phytochem., 197A, 13, 103-105. 

Hasegawa, S. Brewster, L. C. Maier, V. P. Use of limonoate dehydrogenase of Arthrobacter globiformis for the prevention or removal of limonin bitterness in citrus products. J. Food Sci., 1973, 38, 1153-1155. 

Brewster, L. C. Hasegawa, S. Maier, V. P. Bitterness prevention in citrus juices. Comparative activities and stabilities of the limonoate dehydrogenase from Pseudomonas and Arthrobacter. J. Agric. Food Chem., 1976, 2A, 21-2A. 

Specific target enzymes Based on the metabolic and catabolic pathways of limonin in Citrus, several enzymes have been identified for possible genetic manipulation. Specifically, we are currently looking at three enzyme systems 1) limonoate dehydrogenase, 2) limonin UDP-D-glucose transferase and 3) nomilin deacetylase. 

Limonoate dehydrogenase was purified from cell-free extracts of Arthrobacter globiformis by ammonium sulfate fractionation. Blue dye ligand affinity column chromatography and DEAR ion exchange HPLC (29). These steps provided a 428-fold increase in purity over the cell-free extract. The largest increase in purification... 

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