Ragaventhan SANMUGAVELAN, Teow Chong TEOH, Nurnadiah ROSLAN, Zulqarnain MOHAMED

In vitro and in silico studies of chalcone synthase variant 2 in Boesenbergia rotunda and its substrate specificity

In this study, transformation of BrCHS var 2 into B. rotunda cell suspension culture, followed by chalcone synthase enzymaticassay and HPLC analysis was conducted to investigate whether the substrate specificity for BrCHS var 2 is either cinnamoyl-CoAor p-coumaroyl-CoA. The HPLC profile showed an increase in the amount of pinocembrin chalcone when cinnamoyl-CoA andmalonyl-CoA were added but not p-coumaroyl-CoA. Molecular docking was performed to explore the binding of cinnamoyl-CoA andp-coumaroyl-CoA to BrCHS var 2 receptor and the docking results showed that cinnamoyl-CoA formed numerous hydrogen bondsand more negative docked energy than p-coumaroyl-CoA. Cinnamoyl-CoA showed good interactions with Cys 164 to initiate thesubsequent formation of pinocembrin chalcone, whereas the hydroxyl group of p-coumaroyl-CoA formed an unfavorable interactionwith Gln 161 that caused steric hindrance to subsequent formation of naringenin chalcone. Docked conformation analysis results alsoshowed that malonyl-CoA formed hydrogen bonding with Cys 164, His 303, and Asn 336 residues in BrCHS var 2. The results show thatcinnamoyl-CoA is the preferred substrate for BrCHS var 2.

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Abe T, Morita H, Noma H, Kohno T, Noguchi H, Abe I (2007). Structure function analysis of benzalacetone synthase from Rheum palmatum. Bioorg Med Chem Lett 17: 3161-3166.
Abraham MJ, Murtola T, Schulz R, Páll S, Smith JC, Hess B, Lindahl E (2015). GROMACS: high performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX 1-2: 19-25.
Awasthi P, Jamwal VL, Kapoor N, Rasool S, Bahler J (2016). Homology modeling and docking study of chalcone synthase gene (CfCHS) from Coleus forskohlii. Cogent Biology 2: 1175332.
Carpentier SC, Witters E, Laukens K, Deckers P, Swennen R, Panis B (2005). Preparation of protein extracts from recalcitrant plant tissues: an evaluation of different methods for two-dimensional gel electrophoresis analysis. Proteomics 5: 2497-2507.
Christensen AB, Gregersen PL, Schröder J, Collinge DB (1998). A chalcone synthase with an unusual substrate preference is expressed in barley leaves in response to UV light and pathogen attack. Plant Mol Biol 37: 849-857.
de Vries SJ, van Dijk M, Bonvin AM (2010). The HADDOCK web server for data-driven biomolecular docking. Nat Prot 5: 883- 897.
Feinbaum RL, Ausubel FM (1988). Transcriptional regulation of the Arabidopsis thaliana chalcone synthase gene. Mol Cell Biol 8: 1985-1992.
Ferrer JL, Jez JM, Bowman ME, Dixon RA, Noel JP (1999). Structure of chalcone synthase and the molecular basis of plant polyketide biosynthesis. Nature Struct Biol 6: 775-784.
Fliegmann J, Schröder G, Schanz S, Britsch L, Schröder J (1992). Molecular analysis of chalcone and dihydropinosylvin synthase from Scots pine (Pinus sylvestris), and differential regulation of these and related enzyme activities in stressed plants. Plant Mol Biol 18: 489-503.
Goto-Yamamoto N, Wan GH, Masaki K, Kobayashi S (2002). Structure and transcription of three chalcone synthase genes of grapevine (Vitis vinifera). Plant Sci 162: 867-872.
Hatayama M, Ono E, Yonekura-Sakakibara K, Tanaka Y, Nishino T, Nakayama T (2006). Biochemical characterization and mutational studies of a chalcone synthase from yellow snapdragon (Antirrhinum majus) flowers. Plant Biotechnol 23: 373-378.
Jez JM, Bowman ME, Noel JP (2002). Expanding the biosynthetic repertoire of plant type III polyketide synthases by altering starter molecule specificity. P Natl Acad Sci USA 99: 5319- 5324.
Jez JM, Noel JP (2000). Mechanism of chalcone synthase. pKa of the catalytic cysteine and the role of the conserved histidine in a plant polyketide synthase. J Biol Chem 275: 39640-39646.
Kiat TS, Pippen R, Yusof R, Ibrahim H, Khalid N, Rahman NA (2006). Inhibitory activity of cyclohexenyl chalcone derivatives and flavonoids of fingerroot, Boesenbergia rotunda (L.), towards dengue-2 virus NS3 protease. Bioorg Med Chem Lett 16: 3337-3340.
Kirana C, Jones GP, Record IR, McIntosh GH (2007). Anticancer properties of panduratin A isolated from Boesenbergia pandurata (Zingiberaceae). J Nat Med 61: 131-137.
Koduri PK, Gordon GS, Barker EI, Colpitts CC, Ashton NW, Suh DY (2010). Genome-wide analysis of the chalcone synthase superfamily genes of Physcomitrella patens. Plant Mol Biol 72: 247-263.
Koo HJ, McDowell ET, Ma X, Greer KA, Kapteyn J, Xie Z, Descour A, Kim H, Yu Y, Kudrna D (2013). Ginger and turmeric expressed sequence tags identify signature genes for rhizome identity and development and the biosynthesis of curcuminoids, gingerols and terpenoids. BMC Plant Biol 13: 27.
Kreuzaler F, Hahlbrock K (1975). Enzymic synthesis of an aromatic ring from acetate units. Partial purification and some properties of flavanone synthase from cell-suspension cultures of Petroselinum hortense. Eur J Biochem 56: 205-213.
Krieger E, Koraimann G, Vriend G (2002). Increasing the precision of comparative models with YASARA NOVA—a selfparameterizing force field. Proteins 47: 393-402.
Laskowski RA, MacArthur MW, Moss DS, Thornton JM (1993). PROCHECK: a program to check the stereochemical quality of protein structures. J Appl Crystallogr 26: 283-291.
Laskowski RA, Swindells MB (2011). LigPlot+: multiple ligand– protein interaction diagrams for drug discovery. J Chem Inf Model 51: 2778-2786.
Lo C, Coolbaugh RC, Nicholson RL (2002). Molecular characterization and in silico expression analysis of a chalcone synthase gene family in Sorghum bicolor. Physiol Mol Plant P 61: 179-188.
Mallika V, Aiswarya G, Gincy PT, Remakanthan A, Soniya EV (2016). Type III polyketide synthase repertoire in Zingiberaceae: computational insights into the sequence, structure and evolution. Dev Genes Evol 226: 269-285.
Mazumdar AB, Chattopadhyay S (2015). Sequencing, de novo assembly, functional annotation and analysis of Phyllanthus amarus leaf transcriptome using the illumina platform. Front Plant Sci 6: 1199.
Morita H, Takahashi Y, Noguchi H, Abe I (2000). Enzymatic formation of unnatural aromatic polyketides by chalcone synthase. Biochem Bioph Res Co 279: 190-195.
Samappito S, Page J, Schmidt J, De-Eknamkul W, Kutchan TM (2002). Molecular characterization of root-specific chalcone synthases from Cassia alata. Planta 216: 64-71.
Shen YF, Chen GH, Lin SH, Lin G (2016). Molecular modeling and docking calculations of 4-acyloxy-biphenyl-4′- N-butylcarbamates as potential inhibitors of human butyrylcholinesterase. Can J Chem 94: 72-77.
Schüz R, Heller W, Hahlbrock K (1983). Substrate specificity of chalcone synthase from Petroselinum hortense: formation of phloroglucinol derivatives from aliphatic substrates. J Biol Chem 258: 6730-6734.
Sohn JH, Han KL, Lee SH, Hwang JK (2005). Protective effects of panduratin A against oxidative damage of tertbutylhydroperoxide in human HepG2 Cells. Biol Pharm Bull 28: 1083-1086.
Sun W, Meng X, Liang L, Jiang W, Huang Y, He J, Hu H, Almqvist J, Gao X, Wang L (2015). Molecular and biochemical analysis of chalcone synthase from Freesia hybrid in flavonoid biosynthetic pathway. PLoS One 10: e0119054.
Tsou LK, Chen CH, Dutschman GE, Cheng YC, Hamilton AD (2012). Blocking HIV-1 entry by a gp120 surface binding inhibitor. Bioorg Med Chem Lett 22: 3358-3361.
Tuteja JH, Clough SJ, Chan WC, Vodkin LO (2004). Tissue-specific gene silencing mediated by a naturally occurring chalcone synthase gene cluster in Glycine max. Plant Cell 16: 819-835.
Voravuthikunchai SP, Limsuwan S, Supapol O, Subhadhirasakul S (2006). Antibacterial activity of extracts from family zingiberaceae against foodborne pathogens. J Food Safety 26: 325-334.
Wassenaar TA, van Dijk M, Loureiro-Ferreira N, van der Schot G, de Vries SJ, Schmitz C, van der Zwan J, Boelens R, Giachetti A, Ferella L et al. (2012). WeNMR: structural biology on the grid. J Grid Comput 10: 743-767.
Yamazaki Y, Suh DY, Sitthithaworn W, Ishiguro K, Kobayashi Y, Shibuya M, Ebizuka
Y, Sankawa U (2001). Diverse chalcone synthase superfamily enzymes from the most primitive vascular plant, Psilotum nudum. Planta 214: 75-84
Yu B, Zhai H, Wang Y, Zang N, He S, Liu Q (2007). Efficient Agrobacterium tumefaciens-mediated transformation using embryogenic suspension cultures in sweetpotato, Ipomoea batatas (L.) Lam. Plant Cell Tiss Org 90: 265-273.