Identification and characterization of cellular locus of limonin biotransforming enzyme in Pseudomonas putida

Limonin is a highly oxygenated triterpene derivative of class of limonoids which causes delayed bitterness in citrus. Pseudomonas putida (P. putida) is able to reduce 47% and 63% of limonin from juice serum extract and standard limonin, respectively. Biochemical studies with P. putida indicate that probably two metabolic pathways viz. 17-dehydrolimonoid and deoxylimonoid pathway exists in the test organism. Experimental results indicate that the enzyme limonoate dehydrogenase which is found to be localized in periplasmic space of P. putida plays a major role in conversion of limonin to 17-dehydrolimonoate A-ring lactone. Enzymatic studies have shown a 72% reduction in limonin. The experimental results show 9 folds reduction in limonin content in presence of NAD as cofactor. The molecular weight of one of its polypeptide is found to be 66 kDa. The optimum pH and temperature for enzyme activity is 8.5 and 30 degrees C, respectively.