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Isolation, characterization and in silico analysis of 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) gene from Andrographis paniculata (Burm. f) Nees

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Abstract

3-Hydroxy-3-methylglutaryl-coenzymeA reductase (HMGR), the first rate-limiting enzyme of Mevalonate (MVA) pathway was isolated from Andrographis paniculata (ApHMGR) and expressed in bacterial cells. Full length ApHMGR (1937 bp) was submitted to NCBI with accession number MG271748.1. The open reading frame (ORF) was flanked by a 31-bp 5′-UTR, 118-bp 3′-UTR and ApHMGR contained a 1787 bp ORF encoding protein of 595 amino acids. ApHMGR protein was approximately 64 kDa, with isoelectric point of 5.75. Isolated ApHMGR was cloned into pET102 vector and expressed in E. coli BL21 (DE 3) cells, and characterized by SDS-PAGE. HPLC analysis for andrographolide content in leaf, stem and root of A. paniculata revealed highest in leaf tissue. The expression patterns of ApHMGR in different plant tissues using qRT-PCR revealed high in root tissue correlating with HPLC data. Three dimensional (3D) structural model of ApHMGR displayed 90% of the amino acids in most favored regions of the Ramachandran plot with 93% overall quality factor. ApHMGR was highly conserved with plant specific N-terminal membrane domains and C-terminal catalytic regions. Phylogenetic analysis showed A. paniculata sharing common ancestor with Handroanthus impetiginosus. 3D model of ApHMGR was screened for the interaction with substrates NADPH, HMG CoA and inhibitor using Auto Dock Vina. In silico analysis revealed that full length ApHMGR had extensive similarities to other plant HMGRs. The present communication reports the isolation of full length HMGR from A. paniculata, its heterologous expression in bacterial cells and in silico structural and functional characterization providing valuable genomic information for future molecular interventions.

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(Pathway source: Srivatsava and Akhila 2010). (Color figure online)

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References

  1. Singh A, Meena AK, Sudeep Meena, Pant P, Padhi MM (2012) Studies on standardization of Andrographis paniculata nees and identification by HPTLC using andrographolide as marker compound. Int J Phram Pharm Sci. 4:197–200

    Google Scholar 

  2. Akbar S (2012) Andrographis paniculata: a review of pharmacological activities and clinical e!ects. Altern Med Rev 16:66–77

    Google Scholar 

  3. Subramanian R, Asmawi MZ, Sadikun A (2012) A bitter plant with a sweet future? A comprehensive review of an oriental medicinal plant: Andrographis paniculata. Phytochem Rev 11:39–75

    CAS  Google Scholar 

  4. Chao WW, Lin BF (2010) Isolation and identification of bioactive compounds in Andrographis paniculata (Chuanxinlian). Chin Med 5:1–15

    Google Scholar 

  5. Gupta S, Mishra KP, Ganju L (2017) Broad-spectrum antiviral properties of andrographolide. Arch Virol 162:611–623

    CAS  PubMed  Google Scholar 

  6. Nitave SA, Nilesh B, Chougule Koumaravelou (2018) Phytochemical investigation, analgesic and antipyretic activities of ethanolic extract of kariyat. Int J Pharm Sci Res 9:1035–1043

    CAS  Google Scholar 

  7. Suresh V, Raju D, Karuna R et al (2016) Evaluation of antitumor potential of forskolin and andrographolide employing potato tumor bioassay model. Clin Cancer Drugs 3:121–130

    CAS  Google Scholar 

  8. Doi H, Matsui T, Ohye T, Imamura S et al (2018) Andrographolide from Andrographis paniculata shows anti-proliferative activity and anti-inflammatory effects in human monocytic leukemia cells. Fujita Med J 4:55–60

    Google Scholar 

  9. Forestier-Román IS, López-Rivas A, Sánchez-Vázquez MM et al (2019) Andrographolide induces DNA damage in prostate cancer cells. Oncotarget 10:1085–1101

    PubMed  PubMed Central  Google Scholar 

  10. Chassagne F, Haddad M, Amiel A et al (2018) A metabolomic approach to identify anti-hepatocarcinogenic compounds from plants used traditionally in the treatment of liver diseases. Fitoterapia 127:226–236

    CAS  PubMed  Google Scholar 

  11. Khan I, Yusuf MA, Ansari IA, Akhtar MS (2018) Potential of andrographolide, a diterpenoid lactone from Andrographis paniculata: a nature’s treasure for chemoprevention and therapeutics. In: Akhtar MS, Swamy MK (eds) Anticancer plants: mechanisms and molecular interactions, Vol 4. Springer, pp 143–164

  12. Singh S, Pandey P, Ghosh S, Banerjee S (2018) Anti-cancer labdane diterpenoids from adventitious roots of Andrographis paniculata: augmentation of production prospect endowed with pathway gene expression. Protoplasma 255:1387–1400

    CAS  PubMed  Google Scholar 

  13. Rahman UM, Ayoob I, Rehman US et al (2018) Microwave-assisted synthesis of andrographolide analogues as potent β-Glycosidase inhibitors. SynOpen 2:200–206

    Google Scholar 

  14. Gao Z, Yu C, Liang H et al (2018) Andrographolide derivative CX-10 ameliorates dextran sulphate sodium-induced ulcerative colitis in mice: involvement of NF-κB and MAPK signalling pathways. Int Immunopharmacol 57:82–90

    CAS  PubMed  Google Scholar 

  15. Liu Y, Liang RM, Ma QP et al (2017) Synthesis of thioether andrographolide derivatives and their inhibitory effect against cancer cells. Med Chem Comm 8:1268–1274

    CAS  Google Scholar 

  16. Kandanur SG, Kundu S, Cadena C, San Juan H, Bajaj A, Guzman JD, Nanduri S, Golakoti NR (2019) Design, synthesis, and biological evaluation of new 12-substituted-14-deoxy-andrographolide derivatives as apoptosis inducers. Chem Pap 73:1669–1675

    CAS  Google Scholar 

  17. Lee AR, Kwon M, Kang MK, Kim J, Kim SU, Ro DK (2019) Increased sesqui-and triterpene production by co-expression of HMG-CoA reductase and biotin carboxyl carrier protein in tobacco (Nicotiana benthamiana). Metab Eng 52:20–28

    CAS  PubMed  Google Scholar 

  18. Yin JL, Wong WS (2019) Production of santalenes and bergamotene in Nicotiana tabacum plants. PLoS ONE 14:e0203249

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Bansal S, Narnoliya LK, Mishra B et al (2018) HMG-CoA reductase from Camphor Tulsi (Ocimum kilimandscharicum) regulated MVA dependent biosynthesis of diverse terpenoids in homologous and heterologous plant systems. Sci Rep 8:1–15

    Google Scholar 

  20. Jayashree R, Nazeem PA, Rekha K et al (2018) Over-expression of 3-hydroxy-3- methylglutaryl-coenzyme A reductase 1 (hmgr1) gene under super-promoter for enhanced latex biosynthesis in rubber tree (Hevea brasiliensis Muell. Arg.). Plant Physiol Biochem 127:414–424

    CAS  PubMed  Google Scholar 

  21. Yang R, Yuan B, Li W et al (2018) Improving the accumulation of 18 α -and 18 β -glycyrrhizins by over-expressing GuHMGR, GuSQS 1, and GuBAS genes in Glycyrrhiza uralensis. J Tradit Chin Med Sci 4:336–349

    Google Scholar 

  22. Mahobia A, Jha Z (2018) Docking studies of andrographolide and its in vitro validation. Int J Chem Stud 6:1164–1166

    Google Scholar 

  23. Sharma E, Pandey S, Gaur AK (2016) In silico characterization and differential expression pattern analysis of conserved HMG CoA reductase domain isolated from Aconitum balfourii Stapf. 3 Biotech 6:1–8

    Google Scholar 

  24. Bindu BB, Srinath M, Shailaja A, Giri CC (2017) Comparative protein profile studies and in silico structural/functional analysis of HMGR (ApHMGR) in Andrographis paniculata (Burm. f.) Wall. ex Nees. Ann Phytomed 6:30–44

    CAS  Google Scholar 

  25. Srinath M, Shailaja A, Bindu BB, Giri CC (2017) Characterization of 1-deoxy-D-xylulose 5-phosphate synthase (DXS) protein in Andrographis paniculata (Burm. f.) Wall. ex. Nees: A in silico appraisal. Ann Phytomed Int J 6:63–73

    CAS  Google Scholar 

  26. Shailaja A, Bindu BB, Srinath M, Giri CC (2018) In silico structural and functional analysis of copalyl diphosphate synthase enzyme in Andrographis paniculata (Burm. f.) Wall. ex Nees: a plant of immense pharmaceutical value. Ann Phytomed Int J 7:69–77

    CAS  Google Scholar 

  27. Cui Y, Wang Y, Ouyang X, Han Y, Zhu H, Chen Q (2009) Fingerprint profile of active components for Andrographis paniculata Nees by HPLC-DAD. Sens Instrum Food Qual Saf 3:165–179

    Google Scholar 

  28. Garg A, Agrawal L, Misra RC, Sharma S, Ghosh S (2015) Andrographis paniculata transcriptome provides molecular insights into tissue-specific accumulation of medicinal diterpenes. BMC Genomics 16:659

    PubMed  PubMed Central  Google Scholar 

  29. Mou W, Li D, Bu J, Jiang Y, Khan ZU, Luo Z, Mao L, Ying T (2016) Comprehensive analysis of ABA effects on ethylene biosynthesis and signaling during tomato fruit ripening. PLoS ONE 11:1–30

    Google Scholar 

  30. Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497

    CAS  Google Scholar 

  31. Chojnacki S, Cowley A, Lee J, Foix A, Lopez R (2017) Programmatic access to bioinformatics tools from EMBL-EBI update: 2017. Nucleic Acids Res 45:W550–W553

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Koressaar T, Remm M (2007) Enhancements and modifications of primer design program Primer 3. Bioinformatics 23:1289–1291

    CAS  PubMed  Google Scholar 

  33. Untergasser A, Cutcutache I, Koressaar T et al (2012) Primer3 - new capabilities and interfaces. Nucleic Acids Res 40:e115

    CAS  PubMed  PubMed Central  Google Scholar 

  34. San Millán RM, Martínez-Ballesteros I, Rementeria A et al (2013) Online exercise for the design and simulation of PCR and PCR-RFLP experiments. BMC Res Notes 6:513

    PubMed  PubMed Central  Google Scholar 

  35. Waterhouse A, Bertoni M, Bienert S et al (2018) SWISS-MODEL: homology modelling of protein structures and complexes. Nucleic Acids Res 46:W296–W303

    CAS  PubMed  PubMed Central  Google Scholar 

  36. Roy A, Yang J, Zhang Y (2012) COFACTOR: an accurate comparative algorithm for structure-based protein function annotation. Nucleic Acids Res 40:471–477

    Google Scholar 

  37. Watson JD, Sanderson S, Ezersky A et al (2007) Towards fully automated structure-based function prediction in structural genomics: a case study. J Mol Biol 367:1511–1522

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Timothy L. Bailey and Charles Elkan (1994) Fitting a mixture model by expectation maximization to discover motifs in biopolymers”, Proceedings of the Second International Conference on Intelligent Systems for Molecular Biology, AAAI Press, Menlo Park, California, pp.28-36

  39. Marchler-Bauer A, Bo Y, Han L et al (2017) CDD/SPARCLE: functional classification of proteins via subfamily domain architectures. Nucleic Acids Res 45:D200–D203

    CAS  PubMed  Google Scholar 

  40. Robert X, Gouet P (2014) Deciphering key features in protein structures with the new ENDscript server. Nucleic Acids Res 42:W320–W324

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Wass MN, Kelley LA, Sternberg MJ (2010) 3DLigandSite: predicting ligand-binding sites using similar structures. Nucleic Acid Res 38:W469–W473

    CAS  PubMed  PubMed Central  Google Scholar 

  42. Trott O, Olson AJ (2010) AutoDock Vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem 31:455–461

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Boratyn GM, Schäffer AA, Agarwala R et al (2012) Domain enhanced lookup time accelerated BLAST. Biol Direct 7:12

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Huelsenbeck JP, Ronquist F (2001) MRBAYES: Bayesian inference of phylogenetic trees. Bioinformatics 17:754–755

    CAS  PubMed  Google Scholar 

  45. Dereeper A, Guignon V, Blanc G et al (2008) Phylogeny. fr: robust phylogenetic analysis for the non-specialist. Nucleic Acids Res 36:W465–W469

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Dereeper A, Audic S, Claverie JM, Blanc G (2010) BLAST-EXPLORER helps you building datasets for phylogenetic analysis. BMC Evol Biol 12(10):8

    Google Scholar 

  47. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method. Methods 25:402–408

    CAS  PubMed  Google Scholar 

  48. Zhao Q, Li R, Chen X, Yang Q, Li J (2018) Cloning and characterization of the gene encoding 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase from Fritillaria cirrhosa D Don. Braz Arch Biol Technol 61:1–12

    Google Scholar 

  49. Devi K, Patar L, Modi MK, Sen P (2017) An insight into structure, sunction, and expression analysis of 3-hydroxy-3-methylglutaryl-CoA reductase of Cymbopogon winterianus. Bioinform Biol Insights. https://doi.org/10.1177/1177932217701735

    Article  PubMed  PubMed Central  Google Scholar 

  50. Kalita R, Patar L, Shasany AK et al (2015) Molecular cloning, characterization and expression analysis of 3-hydroxy-3-methylglutaryl coenzyme A reductase gene from Centella asiatica L. Mol Biol Rep 42:1431–1439

    CAS  PubMed  Google Scholar 

  51. Ferrero S, Grados-Torrez RE, Leivar P et al (2015) Proliferation and morphogenesis of the endoplasmic reticulum driven by the membrane domain of 3-hydroxy-3-methylglutaryl coenzyme A reductase in plant cells. Plant Physiol 168:899–914

    PubMed  PubMed Central  Google Scholar 

  52. Gu W, Geng C, Xue W et al (2015) Characterization and function of the 3-hydroxy-3-methylglutaryl-CoA reductase gene in Alisma orientale (Sam.) Juz. and its relationship with protostane triterpene production. Plant Physiol Biochem 97:378–389

    CAS  PubMed  Google Scholar 

  53. Tiski I, Marraccini P, Pot D et al (2011) Characterization and expression of two cdna encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase isoforms in Coffee (Coffea arabica L.). Omi A J Integr Biol 15:719–727

    CAS  Google Scholar 

  54. Robertlee J, Kobayashi K, Suzuki M, Muranaka T (2017) AKIN10, a representative Arabidopsis SNF1-related protein kinase 1 (SnRK1), phosphorylates and downregulates plant HMG-CoA reductase. FEBS Lett 591:1159–1166

    CAS  PubMed  Google Scholar 

  55. Srivastava N, Akhila A (2010) Biosynthesis of andrographolide in Andrographis paniculata. Phytochemistry 71:1298-304.

    CAS  PubMed  Google Scholar 

  56. Zeidabadi DD, Javaran MJ, Dehghani H et al (2018) An investigation of the HMGR gene and IPI gene expression in black caraway (Bunium persicum). 3 Biotech 8:405–413

    Google Scholar 

  57. Zhang G, Wu Y, Muhammad ZU, Yang Y, Yu J, Zhang J, Yang D (2019) cDNA cloning, prokaryotic expression and functional analysis of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) in Pogostemon cablin. Protein Expr Purif 163:105454

    CAS  PubMed  Google Scholar 

  58. Rao S, Meng X, Liao Y, Yu T, Cao J, Tan J, Xu F, Cheng S (2019) Characterization and functional analysis of two novel 3-hydroxy-3-methylglutaryl-coenzyme A reductase genes (GbHMGR2 and GbHMGR3) from Ginkgo biloba. Sci Rep 9:1–3

    Google Scholar 

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Acknowledgements

Authors would like to thank DST-PURSE-II sponsored by Department of Science and Technology (DST), and OU-UGC-CPEPA, UGC-BSR-RFSMS sponsored by University Grants Commission (UGC) New Delhi for financial support and fellowship to MS.

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  1. Centre for Plant Molecular Biology (CPMB), Osmania University, Hyderabad, Telangana, 500007, India

    Mote Srinath, Byreddi Bhavani Venkata Bindu, Ayeti Shailaja & Charu Chandra Giri

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  1. Mote Srinath
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Srinath, M., Bindu, B.B.V., Shailaja, A. et al. Isolation, characterization and in silico analysis of 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) gene from Andrographis paniculata (Burm. f) Nees. Mol Biol Rep 47, 639–654 (2020). https://doi.org/10.1007/s11033-019-05172-0

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