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Function(s) and Biological Relevance

Benzylisoquinoline alkaloids (BIAs), plant-specialized metabolites, is the organism that forms TNMT. Tetrahydroprotoberberine N-methyltransferase catalyzes the N-methylation of (S)-stylopine in the pathway leading to protopines and benzo [c] phenanthridines. Also acts on (S)-canadine in a different pathway that leads to the production of phthalideisoquinolines. The substrate specificity of TNMT enzymes appears to arise from the arrangement of subgroup-specific stereospecific recognition elements relative to catalytic elements that are more widely-conserved.

Broader Implications

N-methylation is a recurring feature in the biosynthesis of many plant specialized metabolites, including alkaloids. A crucial step in the conserved central pathway that provides intermediates for the biosynthesis of benzylisoquinoline alkaloids (BIAs). Prominent compounds include the narcotic analgesic morphine, the cough suppressant codeine, the muscle relaxants papaverine and (+)‐tubocurarine, the anti‐microbial agent sanguinarine, and the cholesterol‐lowering drug berberine. In this study, the stereoselectivity of the yellow horned poppy’s enzyme controls what substrates can interact, the products you will get, and how much medicinal compound you can extract from the plant.

Structural highlights and structure-function relationships

GfTNMT was co-crystallized with the cofactor S-adenosyl-L-methionine (dmin = 1.6 A), product S-adenosyl-L-homocysteine (dmin = 1.8 A), or in complex with S-adenosyl-L-homocysteine and (S)-cis-N-methylstylopine (dmin = 1.8 A), These structures reveal for the first time how a mostly L-shaped substrate recognition pocket selects for the (S)-cis configuration of the two central six-membered rings in protoberberine BIA compounds. The substrate specificity of TNMT enzymes appears to arise from the of subgroup-specific stereospecific recognition elements relative to catalytic elements that are more widely conserved among all BIA NMTs. It's catalytic triad consists of His-208, Glu-204, and Glu-207. The ... The ... The binding mode of protoberberine compounds to GfTNMT appears to be similar to coclaurine NMT, with the isoquinoline rings buried deepest in the binding pocket. This binding mode differs from that of pavine NMT, in which the benzyl ring is bound more deeply than the isoquinoline rings. The insights into and catalysis provided here form a sound basis for the rational engineering of for chemoenzymatic synthesis and metabolic engineering.

Energy Transformation

Full-length cDNAs for the three SOMT candidates were cloned into the pRSETA expression vectorwithan N-terminal His6 tag translational fusion. Recombinant SOMT1, SOMT2, and SOMT3 were purified from total protein extract using a cobalt-affinity resin. The recombinant enzymes displayed molecular mass values of approximately 43, 40, and 39 kD as determined by SDS-PAGE, which were marginally higher than the predicted values owing to the N-terminal peptide fusion. The enzyme has a pH optimum of 8.9, a temperature optimum at 40° and a Mr of about 78 000 ± 10%. The Km for (S)-canadine was determined to be 6.4,μM, for (S)-stylopine 3.1 μM and for SAM 12,μM. The enzyme is inhibited by S-adenosyl-l-homocysteine (SAH with a Ki of 24 μM. This is a sample scene created with SAT to by Group, and another to make of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.