title: Production of Δ9-tetrahydrocannabinolic acid from cannabigerolic acid by whole cells of Pichia (Komagataella) pastoris expressing Δ9-tetrahydrocannabinolic acid synthase from Cannabis sativa L.
abstract: OBJECTIVE
  The Δ9-tetrahydrocannabinolic acid synthase (THCAS) from Cannabis sativa was expressed intracellularly in different organisms to investigate the potential of a biotechnological production of Δ9-tetrahydrocannabinolic acid (THCA) using whole cells.
  
  
  RESULTS
  Functional expression of THCAS was obtained in Saccharomyces cerevisiae and Pichia (Komagataella) pastoris using a signal peptide from the vacuolar protease, proteinase A. No functional expression was achieved in Escherichia coli. The highest volumetric activities obtained were 98 pkat ml(-1) (intracellular) and 44 pkat ml(-1) (extracellular) after 192 h of cultivation at 15 °C using P. pastoris cells. Low solubility of CBGA prevents the THCAS application in aqueous cell-free systems, thus whole cells were used for a bioconversion of cannabigerolic acid (CBGA) to THCA. Finally, 1 mM (0.36 g THCA l(-1)) THCA could be produced by 10.5 gCDW l(-1) before enzyme activity was lost.
  
  
  CONCLUSIONS
  Whole cells of P. pastoris offer the capability of synthesizing pharmaceutical THCA production.
authors.affiliation: Laboratory of Technical Biochemistry, Department of Biochemical & Chemical Engineering, TU Dortmund University, Emil-Figge-Str. 66, 44227, Dortmund, Germany.
authors.forename: Bastian
authors.initials: B
authors.lastname: Zirpel
authors.forename: Felix
authors.initials: F
authors.lastname: Stehle
authors.forename: Oliver
authors.initials: O
authors.lastname: Kayser
mesh_terms: D001565:Benzoates / Q000378:metabolism*; D053898:Biosynthetic Pathways; D002188:Cannabis / Q000201:enzymology* / Q000235:genetics; D002474:Cell-Free System; D013759:Dronabinol / Q000378:metabolism*; D019746:Intramolecular Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D010843:Pichia / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D001565:Benzoates; D010940:Plant Proteins; C100679:cannabigerolic acid; D013759:Dronabinol; D019746:Intramolecular Oxidoreductases; C100678:cannabidiolic acid synthase