Lactone hydrolase

Limonin D-ring lactone hydrolase has been isolated from... 

Metabolism in Citrus. Recently, Hasegawa et al. (6) have established the presence of a deoxylimonoid pathway in Citrus. Methyl-14c VIII was metabolized in leaves of calamondin to form a deoxy derivative, showing the presence of epoxidase activity which is required for the first step of the deoxylimonoid pathway. Compound XVIII, the product of the second step, was also isolated from grapefruit seeds (6). Furthermore, deoxylimonate A-ring lactone hydrolase, which is involved in the third step of the pathway, was also detected in grapefruit seeds (6). These findings clearly show that limonoids are metabolized in Citrus not only via the 17-dehydrolimonoid pathway as previously established (22), but also via the deoxylimonoid pathway. 

Limonin D-ring lactone hydrolase is the only limonoid enzyme which has been isolated from Citrus and characterized (11). It is of interest to note that this enzyme is extremely heat resistant. It requires 15 min of heating at 100°C to inactivate it completely. Its functional characteristics are very similar to those of the bacterial hydrolase mentioned previously. 

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. 

Maier, V. P. Hasegawa, S., Hera, E. Limonin D-ring lactone hydrolase. A new enzyme from citrus seeds. Phytochem., 1969, 8, A05-A07. 

A. Hasegawa, S. Metabolism of limonoids Limonin D-ring lactone hydrolase activity in Pseudomonas. J. Agri. Food Chem., 1976, 2A, 2A-26. 

Since the isolation of the oxygenating enzyme and recycling of the cofactor is still difficult for large-scale preparations, the use of whole microorganisms would seem to be advantageous. Whole cells of Acinetobacter are well suited for enantioselective Baeyer-Villiger oxidation of cyclic ketones when a lactone hydrolase inhibitor is used or a hydrolase deficient mutant is... 

Paraxonases (PONs) (52) are a family of mammalian lactone hydrolases, expressed in liver and various tissues, and they can... 

The intact fruits do not contain bitter limonin, but rather a nonbitter precursor, limonoate A-ring lactone (8). When juice is extracted, this nonbitter precursor is gradually converted to limonin under acidic conditions and the conversion is accelerated by the action of limonin D-ring lactone hydrolase, which has been isolated from citrus (9). The bitterness due to nomilin in juices most likely develops in a manner similar to that of limonin bitterness, but this has not yet been directly proven. However, the contribution of nomilin to juices is minor. It occurs mainly In grapefruit juices (4). 

All these degradation pathways were shown to involve a Baeyer-Villiger oxidation of a cycloalkanone that led to formation of the corresponding lactone. Further degradation then occured via hydrolysis of this lactone by a lactone hydrolase which has, in some cases, been isolated. As an example, the reaction sequence for the degradation of cyclopentanol by Pseudomonas sp. NCIMB 98721351 is shown in Fig. 16.5-7. 

The oxidation of racemic fenchone by a Corynebacterium sp.[7<>1 (reclassified as Mycobacterium rhodochrous), an organism which grows at the expense of either (+)- or (-)- camphor, has also been reported. This was shown to lead, in a 45% yield, to a 90/10 mixture of 1,2 and 2,3-fencholides, as shown in Fig. 16.5-12. This result contrasts with the chemical oxidation of fenchone with peracetic acid, where 2,3-fencholide is the major product in a 40/60 mixture. Accumulation of these lactones is a priori surprising as compared with the total degradation of the structurally similar camphor substrate. However this may simply be due to the fact that this lactone, unlike that formed from camphor, is chemically stable in the medium. Of course, one has also to assume that, here again, the strain is devoid of any lactone hydrolase. This bioconversion was the first gram-scale preparative report... 

Development of mutant strains lacking lactone-hydrolases or... 

Fig. 13 Reaction sequence for the conversion of vanillic acid in recombinant cells from Pseudo monas putida. Abbreviations VanAB vanillate O-demethylase, PcaHG dioxygenase, PcaB 3-carboxy-cis,cis-muconate cycloisomerase, PcaC 4-carboxymuconolactone decarboxylase, PcaD p-ketoadipate enol-lactone hydrolase...

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