Structural studies of deoxypodophyllotoxin synthase (DPS), a C-C bond forming enzyme Zoe Ingold 1 , Professor Gideon Grogan 1 , Dr Benjamin Lichman 2 1 York Structural Biology Lab, University of York,UK, 2 Centre for Novel Agricultural Products, University of York, UK https://api.eventsforce.net/rsc/api/v2/events Deoxypodophyllotoxin synthase (DPS) is a member of the 2-oxoglutarate dependent dioxygenase superfamily that performs the ring closing, C-C bond forming reaction shown in Figure 1 . The product of this reaction, deoxypodophyllotoxin, is a polyphenol lignin ultimately derived from tyrosine via the phenylpropanoid pathway. The enzyme was discovered in 2015 during an investigation into the biosynthesis of podophyllotoxin in Podophyllum hexandrum (mayapple), but podophyllotoxin can also be found in several other plants. 1 It is used in the clinic as a topical antiviral as well as being a precursor to several topoisomerase II inhibitors. The enzyme is also under investigation for its industrial potential. 1,2 Carbon-carbon bond formation is of particular interest to biocatalyst development as it is a core aspect of organic chemistry and therefore fine chemical production. Members of the 2-ODD superfamily are already used in the industrial production of antibiotics and modifying DPS will allow the synthesis of deoxypodophyllotoxin derivatives, which may have alternative bioactivities.
Figure 1: DPS catalyses the penultimate reaction, a C-C bond forming and stereospecific ring closure, in the biosyntheticroute for the production of podophyllotoxin from coniferyl alcohol. It is one of two Fe(II) dependent enzymes in the 2-oxoglutarate dependent dioxygenase superfamily known to catalyse the formation of a C-C bond. Previous work on DPS has demonstrated that the enzyme has some degree of substrate promiscuity. 3 In this poster I will present the structure of the enzyme, shown in Figure 2 , and docking studies with the natural product. The structure was solved using X-ray crystallography to a resolution of 1.41 Å. The crystals were obtained in the space group P 2 1 with a single monomer in the asymmetric unit displaying the squashed β-barrel fold common to the 2-ODD superfamily. The open end of this supports the active site with the iron centre coordinated by the “facial triad” of H184, D186 and H239. AutoDock Vina was used to conduct docking studies to determine how the natural substrate (yatein) and product (deoxypodophyllotoxin) may bind. Residues for mutation were selected with the aim of gathering further information about the enzyme mechanism.
Figure 2: Overall structure of DPS, solved to 1.41Å using X-ray crystallography. The structure is colour coded to differentiate between beta sheets (blue) and alpha helices(green), with the facial triadresidues in the active site highlighted
References 1. Lau, W. & Sattely, E. S. Science , 2015 , 349, 1224–1228. 2. Li, J., Zhang, X. & Renata, H. Angewandte Chemie International Edition 2019 , 58, 11657–11660. 3. Lazzarotto, M. et al Angewandte Chemie - International Edition 2019 , 58, 8226–8230.
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