P-Diketone hydrolase

The preferred route for reducing the molecular weight of PVA involves chain scission at the 1,3-diketone site (see Fig. 6). As the diketone element is chemically not very stable, a spontaneous degradation of oxidised PVA was also discussed [80]. Nevertheless, the preferred degradation pathway is most likely the biochemical process because enzymes were identified that showed high activity with diketone substrates [81], especially with oxidised PVA. The p-diketone hydrolase (BDH EC 3.7.1.7) hydrolyses aliphatic p-diketones to form methyl ketones and carboxylic acids in equimolar amounts [82]. The enzymatic cleavage of C-C bonds in p-diketones is not well studied [83]. BDH enzymes could be isolated from different PVA-degrading strains, purified, characterised and cloned [84]. 

In a further example, a biocatalytic route for the production of optically pure 3-substituted cyclohexylamine derivatives from prochiral bicychc P-diketones was established by employing three biocatalytic reaction steps (Scheme 4.16) [53]. The sequence combined the stereoselective hydrolysis of a C-C bond catalyzed by a P-diketone hydrolase [54] (6-oxocamphor hydrolase (OCH) from Rhodococcus sp. [55]), followed by an Upase-catalyzed esterification [Candida antarctica lipase B (CAL-B), Novozyme 435], and a subsequent asymmetric amination by either an (S)-or (1 )-selective m-TA [V.fluvialis [27] or a variant of the Arthrobacter sp. TA [16a] (ArRmutll)]. 

Figure 7.12 Proposal pathway for PVA degradation by various enyme systems. 1) Pyrroloquinoline quinine (PQQ)-dependent PVA hydrogenase 2) oxidised PVA hydrolase 3) PVA-degrading enzyme with dehydrogenase and aldolase activities 4) secondary alcohol oxidase 5) p-diketone hydrolase and 6) PVA-degrading enzyme with oxidase and hydrolase activity. Reproduced with permission from M. Shimao, Current Opinion in Biotechnology, 2001, 12, 3, 242.

Hydrolases were also successfully coupled with co-TAs for the production of 3-cyclohexylamine derivatives from prochiral bicyclic diketones (Scheme 2.17) [41]. The reaction sequence was based on three biocatalytic steps— namely, stereoselective hydrolysis of a C—C bond using a p-diketone hydrolase (6-oxocamphor hydrolase (OCH) from Rhodococcus sp.), followed by Candida antarctica lipase B (C AL-B)-catalyzed esterification, and finally asymmetric amination by stereocomplementary (R)-selective ArRmutll or (S)-selective VF-co-TA. The first two reactions were performed simultaneously in an organic solvent (diisopropyl etirer H O MeOH 97.5/2.5/1). The ami-nation step could also be conducted in an organic solvent after removing the hydrolases by filtration, avoiding the change of the reaction media. 

Chain degradation proceeds at the oxidised sites, where two different routes can be discerned. One way uses the p-hydroxy ketone functionalities introduced by the initial enzymatic attack as direct substrates in an aldolase-like scission reaction. The other way proceeds via further oxidation of this structure to a 1,3-diketone, which serves as a target structure for a hydrolase-type splitting enzyme. 

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