LIMONOATE DEHYDROGENASE FROM ARTHROBACTER-GLOBIFORMIS - THE NATIVE ENZYME AND ITS N-TERMINAL SEQUENCE

Bitter limonoids in citrus juice lower the quality and value of commer cial juices. Limonoate dehydrogenase converts the precursor of bitter limonin, limonoate A-ring lactone, to nonbitter 17-dehydrolimonoate A- ring lactone. This enzyme was isolated from Arthrobacter globiformis c ells by a combination of ammonium sulfate fractionation, Cibacron Blue affinity chromatography and DEAE ion exchange HPLC. Using this protoc ol a 428-fold purification of the enzyme was obtained. Gel filtration HPLC indicated a M, of 118000 for the native enzyme. SDS-PAGE indicate d an individual subunit M, of 31000. N-Terminal sequencing of the prot ein provided a sequence of the first 16 amino acid residues. Since LDH activity in citrus is very low, cloning the gene for this bacterial e nzyme into citrus trees should enhance the natural debittering mechani sm in citrus fruit.