Salutaridinol

Salutaridinol 7-0-acetyltransferase catalyzes the conversion of the phenanthrene alkaloid salutaridinol to salutaridinol-7-Oacetate, the immediate precursor of thebaine along the morphine biosynthetic pathway in P. somniferum (Fig. 10.7).26 Acetyl CoA-dependent acetyltransferases have an important role in plant alkaloid metabolism. They are involved in the synthesis of monoterpenoid indole alkaloids in medicinal plant species such as Rauwolfia serpentina. In this plant, the enzyme vinorine synthase transfers an acetyl group from acetyl CoA to 16-epi-vellosimine to form vinorine. This acetyl transfer is accompanied by a concomitant skeletal rearrangement from the sarpagan- to the ajmalan-type (reviewed in2). An acetyl CoA-dependent acetyltransferase also participates in vindoline biosynthesis in Catharanthus roseus, the source of the chemotherapeutic dimeric indole alkaloid vinblastine (reviewed in2). Acetyl CoA deacetylvindoline 4-O-acetyltransferase catalyzes the last step in vindoline biosynthesis. A cDNA encoding acetyl CoA deacetylvindoline 4-0-acetyltransferase was recently successfully isolated.27... 

Salutaridinol 7-O-acetyltransferase was purified to apparent electrophoretic homogeneity from P. somniferum cell suspension cultures and the amino acid sequence of ten endoproteinase Lys-C-generated peptides was determined.28 A comparison of these amino acid sequences with those available in the GenBank/EMBL sequence databases indicated no relevant similarity to known proteins. The first attempt to isolate a cDNA encoding salutaridinol 1-0-... 

The translation of the sequence of the cDNA encoding deacetylvindoline 4-O-acetyltransferase compared to other putative plant acetyltransferases revealed a conserved region near the carboxy terminus of the proteins. This sequence was used to design a degenerate antisense oligodeoxynucleotide primer for PCR. The sense primer was based upon an internal peptide sequence of salutaridinol 7-0-acetyltransferase. This approach finally yielded a partial cDNA that encoded salutaridinol 7-O-acetyltransferase. The full-length clone was obtained by RACE-PCR and was functionally expressed in S. frugiperda Sf9 cells.28 The amino acid sequence of salutaridinol 7-O-acetyltransferase is most similar (37% identity) to that of deacetylvindoline acetyltransferase of C. roseus.27... 

RNA gel blot analysis of several members of the genus Papaver demonstrated that salutaridinol 7-O-acetyltransferase transcript accumulated in three-week-old seedlings of P. orientale and P. bracteatum, though not in P. atlanticum or P. nudicaule % P. orientale accumulates the alternate biosynthetic precursor oripavine, and P. bracteatum accumulates the morphine biosynthetic precursor... 

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., Acetyl coenzyme A salutaridinol 7-O-acetyltransferase from Papaver somniferum plant cell cultures, J. Biol. Chem., 1995, 270, 31091-31096. 

GROTHE, T., LENZ, R KUTCHAN, T.M., Molecular characterization of the salutaridinol 7-O-acetyltransferase involved in morphine biosynthesis in opium poppy Papaver somniferum, J. Biol. Chem. 2001,276, 30717-30723. 

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],... 

Neodihydrothebaine (7), which was known as a synthetic derivative of the-baine (45), has been found in Papaver bracteatum, from which it was extracted as an inseparable mixture with bractazonine (8). The composition of the mixture was determined by GC-MS analysis and 1H-NMR studies and was further confirmed by comparison with an artificial mixture of synthetic neodihydrothebaine (7) and bractazonine (8) (24). Both alkaloids are considered to be biogenetically derived from thebaine (45) or its immediate precursor salutaridinol (103). 

A parallel biogenetic scheme has been suggested by Theuns et al. (24) for the formation of neodihydrothebaine (7) in Papaver bracteatum. In this case the route proceeds through the isomeric salutaridinol (103), which is derived by para-ortho coupling of reticuline (Scheme 29). It has been suggested that salutaridinol (103) (or thebaine) is the precursor of bractazonine (8), but in this case via a proerythrinadienone formed by aryl migration in the dienol-benzene rearrangement (24) (Scheme 30). 

Recent patents cover the conversion of both isomers of salutaridinol (148 R = Me) into thebaine in 80% yield by the action of thionyl chloride, phosphorus pentachloride, or other acid chlorides, followed by decomposition with bases.180,181 N-Acyl-nor-reticulines have been cyclized by thallium tristrifluoroacetate to the iV-acyl-norsalutaridines [149 R = Ac, F3CCO, cyclo-C3H5CO, EtOCO, or PhCH2OCO), which have been reduced to the corresponding salutaridinols (148) these have been cyclized by sodium hydroxide to the related acyl-northebaines.182... 

Reduction of the ethoxycarbonyl compound (149 R = EtOCO) with lithium aluminium hydride yields salutaridinol (148 R = Me).182... 

Conversion of (S)-reticuline to its ( )-epimer is the first committed step in morphinan alkaloid biosynthesis in certain species. 1,2-Dehydroreticuline reductase catalyzes the stereospecific reduction of 1,2-dehydroreticuline to (7 )-reticuline.39 Intramolecular carbon-carbon phenol coupling of (if)-reticuline by the P450-dependent enzyme salutaridine synthase (STS) results in the formation of salutaridine.40 The cytosolic enzyme, salutaridine NADPH 7-oxidoreductase (SOR), found in Papaver bracteatum and P. somniferum, reduces salutaridine to (7S)-salutaridinol.41 Conversion of (7S)-salutaridinol into thebaine requires closure of an oxide bridge between C-4 and C-5 by acetyl coenzyme A salutaridinol-7-0-acetyltransferase (SAT). The enzyme was purified from opium poppy cultures and the corresponding gene recently isolated (Fig.7.2).42,43 In the last steps of morphine... 

The systemic transport of pathway intermediates or end-products might also occur in BA biosynthesis. Several BA biosynthetic enzymes are preferentially active in certain organs. Transcripts encoding various TYDC isoforms were most abundant in roots, or roots and stems.3 NCS was also most active in roots and stems of opium poppy,112 whereas CYP80B1 transcripts were most abundant in stems followed by roots, leaves, and floral tissues.148 STS and SOR, which convert (R)-reticuline to salutaridinol, exhibited abundant activity in roots and shoots.40,41 In contrast, COR... 

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