Biosynthesis of menisporopsin A, a fungal macrocyclic polylactone Pakorn Wattana-Amorn 1 , Thanakorn Deelee 1 , Ashley J. Winter 2 , Passorn Wonnapinij 3 , Ajaraporn Sriboonlert 3 , Colin M. Lazarus 4 , Matthew P. Crump 2 , Paul R. Race 5 1 Department of Chemistry, Kasetsart University, Thailand, 2 School of Chemistry, University of Bristol, UK, 3 Department of Genetics, Kasetsart University, Thailand, 4 School of Biological Sciences, University of Bristol, UK, 5 School of Biochemistry, University of Bristol, UK Menisporopsin A is a bioactive macrocyclic polylactone exhibiting cytotoxic and antimalarial activities. This compound is produced by the fungus, Menisporopsis theobromae BCC 4162, and its biosynthetic pathway has been proposed based on the 13 C labelling experiments. Both reducing (R) and non-reducing (NR) polyketide synthases (PKSs) are expected to catalyze the formation of each menisporopsin A subunit, and a non-ribosomal peptide synthetase (NRPS)-like enzyme is possibly involved in multiple esterifications and cyclolactonization. Transcriptomic data of M. theobromae BCC 4162 during the production phase of menisporopsin A revealed two PKS genes that are possibly involved in the biosynthesis of menisporopsin A. They are an R-PKS gene, men1 , and an NR-PKS gene, men2 , which were cloned into separate vectors for heterologous expression in Aspergillus oryzae NSAR1 . Unexpectedly, coexpression of men1 and men2 are sufficient to catalyze the formation of ascotrichalactone A, a possible intermediate of menisporopsin A. This could be a result of the unusal functions of the thioesterase (TE) and the doublet acyl carrier protein (ACP) domains of Men2. Solution characterization of these domains (ACP1-ACP2-TE and ACP2-TE) performed by small-angle X-ray scattering (SAXS) in combination with homology modelling also revealed the highly flexible interdomain linkers between these domains. These results provide the first structural information of TE and ACP arrangement in the fungal iterative PKS. References 1. Bunnak, W., Winter, A.J., Lazarus, C.M., Crump, M.P., Race, P.R., Wattana-Amorn, P. (2021) SAXS reveals highly flexible interdomain linkers of tandem acyl carrier protein-thioesterase domains from a fungal nonreducing polyketide synthase. FEBS lett . 595, 133-144. 2. Bunnak, W., Wonnapinij, P., Sriboonlert, A., Lazarus, C.M., Wattana-Amorn, P. (2019) Heterologous biosynthesis of a fungal macrocyclic polylactone requires only two iterative polyketide synthases. Org. Biomol. Chem. 17, 374-379. 3. Wattana-Amorn, P., Juthaphan, P., Sirikamonsil, M., Sriboonlert, A., Simpson, T.J., Kongkathip, N. (2013) Biosynthetic Origins of Menisporopsin A. J. Nat. Prod. 76 , 1235-1237.
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