Codeinone reductase

Fig. 10.6 Reaction catalyzed by codeinone reductase (COR), the penultimate step in morphine biosynthesis.

The peptide sequences obtained for codeinone reductase aligned well with the amino acid sequences for 6 -deoxychalcone synthase (chalcone reductase) from alfalfa, Glycerrhiza, and soybean. Knowledge of the relative positions of the peptides allowed for a quick RT-PCR based isolation of cDNAs encoding codeinone reductase from P. somniferum. The codeinone reductase isoforms are 53 % identical to chalcone reductase from soybean.25 By sequence comparison, both codeinone reductase and chalcone reductase belong to the aldo/keto reductase family, a group of structurally and functionally related NADPH-dependent oxidoreductases, and thereby possibly arise from primary metabolism. Six alleles encoding codeinone... 

With a morphine biosynthetic gene in hand, we believed we could begin to address the question why only P. somniferum produces morphine, while other Papaver species such as P. rhoeas, P. orientale, and P. bracteatum do not. Unexpectedly, we found that the codeinone reductase transcript was present to some degree in all four species investigated. A review of the literature revealed no alkaloids reported in P. rhoeas for which codeinone reductase should participate in the synthesis. Similarly, P. orientale accumulates the alternate morphine biosynthetic precursor oripavine, but codeinone reductase is not involved in the biosynthesis of oripavine, acting instead after this alkaloid along the biosynthetic pathway to morphine.22 P. bracteatum produces the morphine precursor thebaine as a major alkaloid. As for oripavine in P. orientale, codeinone reductase would act in P. bracteatum after thebaine formation on the pathway to morphine. It appears, therefore, that the reason that P. rhoeas, P. orientale, and P. bracteatum do not produce morphine is not related to the absence of the transcript of the morphine biosynthesis-specific gene codeinone reductase. The expression of codeinone reductase may simply be an evolutionary remnant in these species. 

Fig. 10.8 Selected cDNAs isolated in recent years that encode enzymes involved in the biosynthesis of various classes of isoquinoline alkaloids. 6-OMT, norcoclaurine 6-0-methyltransferase 23 CYP80A1, berbamunine synthase 19 CYP80B1, (S)-A-methylcoclaurine 3 -hydroxylase 20 CPR, cytochrome P-450 reductase 29 4 -OMT, (5)-3 -hydroxy-A-methylcoclaurine 4 -0-methyltransferase 30 BBE, berberine bridge enzyme 12 SalAT, salutaridinol 7-O-acetyltransferase 28 COR, codeinone reductase.25...

LENZ, R., ZENK, M.H., Purification and properties of codeinone reductase (NADPH) from Papaver somniferum cell cultures and differentiated plants, Eur. J. Biochem., 1995, 233, 132-139. 

UNTERLINNER, B LENZ, R KUTCHAN, T.M., Molecular cloning and functional heterologous expression of codeinone reductase The penultimate enzyme in morphine biosynthesis in the opium poppy Papaver somniferum, Plant J., 1999,18,465-475. 

The isolation of the cDNAs encoding the enzymes involved in diverse isoquinoline alkaloid formation in plants and microorganisms allowed the first metabolic engineering routes to be developed and paved the way for new ways of future production of isoquinoline alkaloids. For instance, transgenic opium poppy plants were created in which codeinone reductase was suppressed by RNAi, resulting in the substitution of morphine synthesis with the non-narcotic precursor reticuline [110]. In a similar approach, RNAi suppression or overexpression of salutaridinol 1-0-acetyltransferase in opium poppy led to accumulation of salutaridine or increase of morphine, codeine and thebaine content [111], suppression of the BBE led to accumulation of berberine in California poppy cells [112],... 

Figure 2.8 Biosynthesis of morphine via the conversion of (S)-reticuline to (R)-reticuline, salutaridine and thebaine. SalS, salutaridine synthase SaIR, salutaridine reductase THS, thebaine synthase COR, codeinone reductase.

Figure 7.17 Phyiogenetic reiationships in key enzymes of iater steps in SM pathways, based on amino acid sequences, (a) Chaicone synthase (CHS), (b) Strictosidine synthase (STS), (c) Berberine bridge enzyme (BBE). (d) Codeinone reductase (CR).

The localization of isoquinoline biosynthesis has been investigated at the cellular level in intact poppy plants by using in situ RNA hybridization and immunoflouresence microscopy. The localization of 4 -OMT (reticuline biosynthesis), berberine bridge enzyme (saguinarine biosynthesis), salutaridinol acetyltransferase (morphine biosynthesis), and codeinone reductase (morphine biosynthesis) has been probed. 4 -OMT and salutaridinol acetyltransferase are localized to parenchyma cells, whereas codeinone reductase is localized to laticifer cells in... 

Lenz R, Zenk MH. Stereoselective reduction of codeinone, the penultimate enzymic step during morphine biosynthesis in Papaver somniferum Tetrahedron Lett. 1995 36 2449-2452. UnterUnner B, Lenz R, Kutchan TM. Molecular cloning and functional expression of codeinone reductase. Plant J. 1999 18 465-475. 

Frick S, Kutchan TM, Fist AJ. Increasing morphinan alkaloid production by overexpressing codeinone reductase in transgenic Papaver somniferum. Plant Biotechnol. J. 2007 5 26-37. 73. 

Codeinone reductase Papaver somnlferum aMo/keto reductase AGOSC... 

Scheme 1. Biosynthesis of the benzylisoquinoline alkaloids berberine, morphine, and sang-uinarine. Enzymes for which corresponding molecular clones have been isolated are shown in bold. Abbreviations 4 OMT, 3 -hydroxy-A-methylcoclaurine 4 -0-methyltransferase 60MT, norcoclaurine 6-0-methyltransferase 70MT, reticuline 7-0-methyltransferase BBE, berberine bridge enzyme CFS, cheilanthifoline synthase CNMT, coclaurine A-methyltransf-erase COR, codeinone reductase CYP719A1, canadine synthase CYP80A1, berbamunine synthase CYP80B1, A-methylcoclaurine 3 -hydroxylase DBOX, dihydrobenzophenanthri-dine oxidase DRR, 1,2-dehydroreticuline reductase DRS, 1,2-dehydro reticuline synthase MSH, A-methylstylopine 14-hydroxylase NCS, norcoclaurine synthase P6H,...

Several studies have reported the production in microorganisms of plant enzymes that are involved in alkaloid biosynthesis [29, 99-102], In this sense, Unterlinner et al. [100] have cloned and expressed in Escherichia coli four cDNAs encoding for different isoforms of the Codeinone reductase NADPH-dependent enzyme isolated from Papaver somniferum. In this report has been investigated the substrate specificity of the enzyme and the structural analysis, being the first report about the cloning and expression of genes of the biosynthetic pathway of morphine [90],... 

As shown in Scheme 13.43, the conversion of thebaine to codeine and thence to morphine is effected by enzymes that have not yet been completely characterized. It is clear that methyl groups must be removed and a reduction [codeinone reductase, EC 1.1.1.247, NAD(P)H NAD(P)] must be effected. The sequence is not clear, although there does seem to be a general consensus that the scheme shown probably applies. 

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