Skip to main content

Comparative analyses of squalene synthase (SQS) proteins in poplar and pine by using bioinformatics tools

Abstract

Squalene synthase (SQS, EC 2.5.1.21) is a major enzyme in biosynthesis of isoprenoid (farnesyl pyrophosphate (FPP) squalene). In the present study, we have analyzed SQS enzymes of black cottonwood (Populus trichocarpa, hereafter Pt) and Masson’s pine (Pinus massoniana, hereafter Pm) using bioinformatics tools. PtSQS and PmSQS sequences were found to have very similar physicochemical properties with “squalene/phytoene synthase” domain structure (PF00494). PtSQS sequence was 47.3 kDa weight and 413 amino acids long with a pI value of 6.86, while PmSQS was 46.6 kDa weight and 409 amino acids long with a pI of 7.92. Alignment of SQS protein sequences in 15 plant species showed a highly similar conserved pattern and included 77DTVED81 and 213DYLED217 motifs, which are rich in aspartic acids, for FPP binding sites. In phylogenetic tree, monocots and polycot were clearly separated from dicots with high bootstrap value (99 %). A total of 10 interaction partners were predicted for PtSQS and PmSQS proteins. Nine of them were hypothetical proteins (related with phytosterol biosynthesis), while one was putative uncharacterized protein. Similar 3D structures and identical binding sites were predicted for pine and poplar. In docking, FPP-PtSQS was found to make 8 H bonds with Asp81, Asp217, Glu80, and Gln206 residues in poplar with highest affinity while FPP-PmSQS made 7 H bonds with Arg49, Arg74, Ser48, and Val47 residues in pine with highest affinity. The results of this study will provide valuable theoretical knowledge for future studies of identification and characterization of SQS genes and proteins in various tree species and will provide an insight for studies of biotechnological manipulation of sterol biosynthesis pathway to enhance the plant stress tolerance and productivity.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

References

  • Akamine S, Nakamori K, Chechetka SA, Banba M, Umehara Y, Kouchi H, Hata S (2003) cDNA cloning, mRNA expression, and mutational analysis of the squalene synthase gene of Lotus japonicas. BBA- Gene Struct Expr 1626(1):97–101

    CAS  Article  Google Scholar 

  • Bailey TL, Boden M, Buske FA, Frith M, Grant CE, Clementi L et al (2009) MEME SUITE: tools for motif discovery and searching. Nucleic Acids Res. doi:10.1093/nar/gkp335

  • Benkert P, Künzli M, Schwede T (2009) QMEAN server for protein model quality estimation. Nucleic Acids Res 37(Web Server issue):W510–4

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Bhat WW, Lattoo SK, Razdan S, Dhar N, Rana S, Dhar RS et al (2012) Molecular cloning, bacterial expression and promoter analysis of squalene synthase from Withania somnifera (L.) Dunal. Gene 499(1):25–36

    CAS  Article  PubMed  Google Scholar 

  • Burleigh JG, Barbazuk WB, Davis JM, Morse AM, Soltis PS (2012) Exploring diversification and genome size evolution in extant gymnosperms through phylogenetic synthesis. J Bot. doi:10.1155/2012/292857

    Google Scholar 

  • Dundas J, Ouyang Z, Tseng J, Binkowski A, Turpaz Y, Liang J (2006) CASTp: computed atlas of surface topography of proteins with structural and topographical mapping of functionally annotated residues. Nucleic Acid Res 34:W116–W118

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Franceschini A, Szklarczyk D, Frankild S, Kuhn M, Simonovic M, Roth A et al (2013) STRING v9. 1: protein-protein interaction networks, with increased coverage and integration. Nucleic Acids Res 41(Database issue):D808–15

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Geourjon C, Deleage G (1995) SOPMA: significant improvements in protein secondary structure prediction by consensus prediction from multiple alignments. Comput Appl Biosci 11:681–684

    CAS  PubMed  Google Scholar 

  • Ginzberg I, Thippeswamy M, Fogelman E, Demirel U, Mweetwa AM, Tokuhisa J et al (2012) Induction of potato steroidal glycoalkaloid biosynthetic pathway by overexpression of cDNA encoding primary metabolism HMG-CoA reductase and squalene synthase. Planta 235(6):1341–1353

    CAS  Article  PubMed  Google Scholar 

  • Grover A, Samuel G, Bisaria VS, Sundar D (2013) Enhanced withanolide production by overexpression of squalene synthase in Withania somnifera. J Biosci Bioeng 115(6):680–685

    CAS  Article  PubMed  Google Scholar 

  • Gu P, Ishii Y, Spencer TA, Shechter I (1998) Function-structure studies and identification of three enzyme domains involved in the catalytic activity in rat hepatic squalene synthase. J Biol Chem 273(20):12515–12525

    CAS  Article  PubMed  Google Scholar 

  • Gupta N, Sharma P, Kumar RS, Vishwakarma RK, Khan BM (2012) Functional characterization and differential expression studies of squalene synthase from Withania somnifera. Mol Biol Rep 39(9):8803–8812

    CAS  Article  PubMed  Google Scholar 

  • Hanley KM, Nicolas O, Donaldson TB, Smith-Monroy C, Robinson GW, Hellmann GM (1996) Molecular cloning, in vitro expression and characterization of a plant squalene synthase cDNA. Plant Mol Biol 30:1139–1151

    CAS  Article  PubMed  Google Scholar 

  • Inoue T, Osumi T, Hata S (1995) Molecular cloning and functional expression of a cDNA for mouse squalene synthase. Biochim Biophys Acta 1260:49–54

    Article  PubMed  Google Scholar 

  • Jennings SM, Tsay YH, Fisch TM, Robinson GW (1991) Molecular cloning and characterization of the yeast gene for squalene synthetase. Proc Natl Acad Sci 88:6038–6042

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Johnson JE, Cornell RB (1999) Amphitropic proteins: regulation by reversible membrane interactions (review). Mol Membr Biol 16(3):217–235

    CAS  Article  PubMed  Google Scholar 

  • Kalra S, Kumar S, Lakhanpal N, Kaur J, Singh K (2013) Characterization of squalene synthase gene from Chlorophytum borivilianum (Sant. and Fernand.). Mol Biotech 54(3):944–953

    CAS  Article  Google Scholar 

  • Kelley LA, Sternber MJ (2009) Protein structure prediction on the Web: a case study using the Phyre server. Nat Protoc 4:363–371

    CAS  Article  PubMed  Google Scholar 

  • Kim TD, Han JY, Huh GH, Choi YE (2011a) Expression and functional characterization of three squalene synthase genes associated with saponin biosynthesis in Panax ginseng. Plant Cell Physiol 52(1):125–137

  • Kim YS, Cho JH, Park S, Han JY, Back K, Choi YE (2011b) Gene regulation patterns in triterpene biosynthetic pathway driven by overexpression of squalene synthase and methyl jasmonate elicitation in Bupleurum falcatum. Planta 233(2):343–355

  • Kribii R, Arró M, Del Arco A, González V, Balcells L, Delourme D, Ferrer A, Karst F, Boronat A (1997) Cloning and characterization of the Arabidopsis thaliana SQS1 gene encoding squalene synthase—involvement of the C-terminal region of the enzyme in the channeling of squalene through the sterol pathway. Eur J Biochem 249:61–69

    CAS  Article  PubMed  Google Scholar 

  • Kuang YW, Sun FF, Wen DZ, Zhou GY, Zhao P (2008) Tree-ring growth patterns of Masson pine (P. massoniana) during the recent decades in the acidification Pearl River Delta of China. For Ecol Manag 255:3534–3540

    Article  Google Scholar 

  • Laule O, Fürholz A, Chang HS, Zhu T, Wang X, Heifetz PB et al (2003) Crosstalk between cytosolic and plastidial pathways of isoprenoid biosynthesis in Arabidopsis thaliana. Proc Natl Acad Sci 100(11):6866–6871

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Lee MH, Jeong JH, Seo JW, Shin CG, Kim YS et al (2004) Enhanced triterpene and phytosterol biosynthesis in Panax ginseng overexpressing squalene synthase gene. Plant Cell Physiol 45:976–984

    CAS  Article  PubMed  Google Scholar 

  • Leitch IJ, Leitch AR (2013) Genome size diversity and evolution in land plants. Springer, Vienna, pp 307–322

    Book  Google Scholar 

  • Lichtenthaler HK (2007) Biosynthesis, accumulation and emission of carotenoids, α- tocopherol, plastoquinone, and isoprene in leaves under high photosynthetic irradiance. Photosynth Res 92(2):163–179

    CAS  Article  PubMed  Google Scholar 

  • Manavalan LP, Chen X, Clarke J, Salmeron J, Nguyen HT (2012) RNAi-mediated disruption of squalene synthase improves drought tolerance and yield in rice. J Exp Bot 63(1):163–175

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Morris GM, Huey R, Lindstrom W, Sanner MF, Belew RK, Goodsell DS, Olson AJ (2009) AutoDock4 and AutoDockTools4: automated docking with selective receptor flexibility. J Comput Chem 30(16):2785–2791

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Nakashima T, Inoue T, Oka A, Nishino T, Osumi T, Hata S (1995) Cloning, expression, and characterization of cDNAs encoding Arabidopsis thaliana squalene synthase. Proc Natl Acad Sci 92:2328–2332

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Nguyen HTM, Neelakadan AK, Quach TN, Valliyodan B, Kumar R, Zhang Z, Nguyen HT (2013) Molecular characterization of Glycine max squalene synthase genes in seed phytosterol biosynthesis. Plant Physiol Bioch 73:23–32

    CAS  Article  Google Scholar 

  • Palta JP, Whitaker BD, Weiss LS (1993) Plasma membrane lipids associated with genetic variability in freezing tolerance and cold acclimation of Solanum species. Plant Physiol 103(3):793–803

    CAS  PubMed  PubMed Central  Google Scholar 

  • Pandit J, Danley DE, Schulte GK, Mazzalupo S, Pauty TA, Hayward CM, Hamanaka ES, Thompson JF, Harwood HJ Jr (2000) Crystal structure of human squalene synthase, a key enzyme in cholesterol biosynthesis. J Biol Chem 275:30610–30617

    CAS  Article  PubMed  Google Scholar 

  • Piironen V, Lindsay DG, Miettinen TA, Toivo J, Lampi AM (2000) Plant sterols: biosynthesis, biological function and their importance to human nutrition. J Sci Food Agric 80:939–966

    CAS  Article  Google Scholar 

  • Sanchita SG, Sharma A (2014) In silico study of binding motifs in squalene synthase enzyme of secondary metabolic pathway solanaceae family. Mol Biol Rep 41:7201–7208

    CAS  Article  PubMed  Google Scholar 

  • Schaller H (2003) The role of sterols in plant growth and development. Prog Lipid Res 42(3):163–175

    CAS  Article  PubMed  Google Scholar 

  • Seo JW, Jeong JH, Shin CG, Lo SC, Han SS et al (2005) Overexpression of squalene synthase in Eleutherococcus senticosus increases phytosterol and triterpene accumulation. Phytochemistry 66:869–877

    CAS  Article  PubMed  Google Scholar 

  • Singh AK, Dwivedi V, Rai A, Pal S, Reddy SGE, Rao DKV et al (2015) Virus‐induced gene silencing of Withania somnifera squalene synthase negatively regulates sterol and defense‐related genes resulting in reduced withanolides and biotic stress tolerance. Plant Biotech J doi:10.1111/pbi.12347

    Google Scholar 

  • Singh G, Sharma A (2014) In silico study of binding motifs in squalene synthase enzyme of secondary metabolic pathway solanaceae family. Mol Biol Rep 41(11):7201–7208

    Article  PubMed  Google Scholar 

  • Sonnhammer EL, Eddy SR, Durbin R (1997) Pfam: a comprehensive database of protein domain families based on seed alignments. Proteins: Struct Funct Genet 28(3):405–420

  • Stamellos KD, Shackelford JE, Shechter I, Jiang G, Conrad D, Keller GA, Krisans SK (1993) Subcellular localization of squalene synthase in rat hepatic cells. Biochemical and immunochemical evidence. J Biol Chem 268:12825–12836

    CAS  PubMed  Google Scholar 

  • Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28(10):2731–2739

  • Tansey TR, Shechter I (2000) Structure and regulation of mammalian squalene synthase. BBA-Mol Cell Biol L 1529(1):49–62

    CAS  Google Scholar 

  • Thuduppathy GR, Craig JW, Kholodenko V, Schon A, Hill RB (2006) Evidence that membrane insertion of the cytosolic domain of Bcl-X L is governed by an electrostatic mechanism. J Mol Biol 359(4):1045–1058

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Tuskan GA, Difazio S, Jansson S, Bohlmann J, Grigoriev I, Hellsten U et al (2006) The genome of black cottonwood, Populus trichocarpa (Torr. & Gray). Sci 313(5793):1596–1604

    CAS  Article  Google Scholar 

  • Uchida H, Yamashita H, Kajikawa M, Ohyama K, Nakayachi O et al (2009) Cloning and characterization of a squalene synthase gene from a petroleum plant, Euphorbia tirucalli L. Planta 229:1243–1252

    CAS  Article  PubMed  Google Scholar 

  • Vishwakarma RK, Patel K, Sonawane P, Kumari U, Singh S, Abbassi S et al. (2015). Squalene Synthase Gene from Medicinal Herb Bacopa monniera: Molecular Characterization, Differential Expression, Comparative Modeling, and Docking Studies. Plant Mol Biol Rep 33(6):1675–1685

  • Wang XQ, Ran JH (2014) Evolution and biogeography of gymnosperms. Mol Phylogenet Evol 75:24–40

    Article  PubMed  Google Scholar 

  • Wang K, Senthil-Kumar M, Ryu CM, Kang L, Mysore KS (2012) Phytosterols play a key role in plant innate immunity against bacterial pathogens by regulating nutrient efflux into the apoplast. Plant Physiol 158:1789–1802

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Willard L, Ranjan A, Zhang H, Monzavi H, Boyko RF, Sykes BD, Wishart DS (2003) VADAR: a web server for quantitative evaluation of protein structure quality. Nucleic Acids Res 31:3316–3319

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Zhao RY, Xiao W, Cheng HL, Zhu P, Cheng KD (2010) Cloning and characterization of squalene synthase gene from Fusarium fujikuroi (Saw.) Wr. J Ind Microbiol Biotechnol 37:1171–1182

    CAS  Article  PubMed  Google Scholar 

  • Zhan DL, Zhang Y, Song YW, Sun H, Li ZS, Han WW, Liu JS (2012) Computational studies of squalene synthase from Panax ginseng: homology modeling, docking study and virtual screening for a new inhibitor. J Theor Comput Chem 11:1101–1120

    CAS  Article  Google Scholar 

  • Zhang LY, Deng XW, Lei XD, Xiang WH, Peng CH, Lei PF, Yan WD (2012) Determining stem biomass of Pinus massoniana L. through variations in basic density. Forestry 85:601–609

    Article  Google Scholar 

  • Zheng Z, Cao X, Li C, Chen Y, Yuan B, Xu Y, Jiang J (2013) Molecular cloning and expression analysis of a squalene synthase gene from a medicinal plant, Euphorbia pekinensis Rupr. Acta Physiol Plant 35:3007–3014

    CAS  Article  Google Scholar 

  • Ye Y, Wang R, Jin L, Shen J, Li X, Yang T et al (2014) Molecular cloning and differential expression analysis of a squalene synthase gene from, an important pharmaceutical plant. Mol Biol Rep 9(41):6097–6104

    Article  Google Scholar 

  • Zook MN, Kuc JA (1991) Induction of sesquiterpene cyclase and suppression of squalene synthetase activity in elicitor-treated or fungal-infected potato tuber tissue. Physiol Mol Plant P 39(5):377–390

    CAS  Article  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Ertugrul Filiz.

Ethics declarations

The authors declared that they have no conflicts of interest.

Additional information

Communicated by J.L. Wegrzyn

Electronic Supplementary Material

Below is the link to the electronic supplementary material.

Table S1

Table S1 (DOCX 21 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Filiz, E., Ozyigit, I.I. & Vatansever, R. Comparative analyses of squalene synthase (SQS) proteins in poplar and pine by using bioinformatics tools. Tree Genetics & Genomes 12, 32 (2016). https://doi.org/10.1007/s11295-016-0992-0

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s11295-016-0992-0

Keywords

  • Phytosterol
  • Molecular docking
  • 3D structure
  • Protein–protein interaction