Genome-wide detection of terpene synthase genes in holy basil (Ocimum sanctum L.)

Yogesh Kumar, Feroz Khan, Shubhra Rastogi, Ajit Kumar Shasany, Tzen-Yuh Chiang
2018 PLoS ONE  
Holy basil (Ocimum sanctum L.) and sweet basil (Ocimum basilicum L.) are the most commonly grown basil species in India for essential oil production and biosynthesis of potentially volatile and non-volatile phytomolecules with commercial significance. The aroma, flavor and pharmaceutical value of Ocimum species is a significance of its essential oil, which contains most of the monoterpenes and sesquiterpenes. A large number of plants have been studied for characterization and identification of
more » ... identification of terpene synthase genes, involved in terpenoids biosynthesis. The goal of this study is to discover and identify the putative functional terpene synthase genes in O. sanctum. HMMER search was performed by using a set of 13 well sequenced and annotated plant genomes including the newly sequenced genome of O. sanctum with Pfam-A database locally, using HMMER 3.0 hmmsearch for the two Pfam domains (PF01397 and PF03936). Using this search method 81 putative terpene synthases genes (OsaTPS) were identified in O. sanctum; the study further reveals 47 OsaTPS were putatively functional genes, 19 partial OsaTPS, and 15 OsaTPS as probably pseudogenes. All these identified OsaTPS genes were compared with other plant species, and phylogenetic analysis reveals the subfamily classification of OsaTPS in TPS-a, -b, -c, -e, -f and TPS-g subfamilies clusters. This genome-wide identification of OsaTPS genes, their phylogenetic analysis and secondary metabolite pathway mapping predictions together provide a comprehensive understanding of the TPS gene family in Ocimum sanctum and offer opportunities for the characterization and functional validation of numbers of terpene synthase genes. OPEN ACCESS Citation: Kumar Y, Khan F, Rastogi S, Shasany AK (2018) Genome-wide detection of terpene synthase genes in holy basil (Ocimum sanctum L.). PLoS ONE 13(11): e0207097. known as isoprenyl diphosphate synthases [5] . The condensation reaction by prenyltransferases produces geranyl diphosphate (GPP C 10 ), farnesyl diphosphate (FPP C 15 ) in the cytosol and geranylgeranyl pyrophosphate (GGPP C 20 ) in the chloroplast. These all prenyl pyrophosphates work as a substrate for the terpene synthases, which further performed the catalysis for the production of different terpenoids: hemiterpene (C 5 ) is produced directly by isopentyl pyrophosphate, monoterpene (C 10 ) from GPP, sesquiterpene (C 15 ) from FPP, diterpene (C 20 ) form GGPP, and other terpenes like sesterterpenes (C 25 ), triterpenes (C 30 ), sesquarterpenes (C 35 ), tetraterpenes (C 40 ) and polyterpene (>C 40 ) [6]. These terpenes are responsible for plant defense against biotic and abiotic stresses, used to attract insects for pollination, and used by human for their beneficial purposes as a natural flavor, for perfumery, and for cosmetics. These terpenoids are synthesized by specialized genes know as terpene synthases. The terpene synthase genes are classified into different classes. According to their genomic structures (including intron and exon numbers), terpene synthase sequences (TPSs) can be classified into three classes Class I, Class II and Class III terpene synthases. The class I terpene synthase genes contain 12-14 introns, and 13-15 exons, class II genes possess nine introns and ten exons. Moreover, class III terpene synthase genes have six introns and seven exons [7] [8] [9] . Based on catalytic mechanism and product formed, TPSs genes are also classified into two classes: class I and class II. The class I and II TPSs have unique conserved amino acid motifs which are essential for catalysis [10, 11] . Class I TPSs has a C-terminal domain (also referred to as α-domain or class I fold) which catalyzes the ionization of substrate (i.e., prenyl diphosphate) mediated by a divalent cation. This metal-dependent ionization of substrate can further lead to cyclizations, hydride shifts, and structural rearrangements to produce the end product. The α-domain present in this class TPSs adopt the α-helical protein fold and holds two metal binding motifs 'DDXXD' a highly conserved and less conserved 'NSE/DTE' positioned on opposing helices near the entrance of the active site. The class II terpene synthases comprise a functional N-terminal (β-domain), with a third "insertion" γ-domain forming a vestigial class II fold. The enzymes of this class contain conserved functional 'DXDD' motif which resides in a separate β-domain and accountable for the protonation-initiated cyclization of the substrate [11, 12] . The γ-domain carries a highly acidic EDXXD like motif, which contributes to the activity of class II TPSs (Fig 1) [13]. Many terpene synthases also have a highly conserved RR (x)8W motif downstream of the N-terminal transit peptide while this motif does not require in monoterpene synthase activity. According to recent subfamilies classification, TPS genes are classified into eight subfamilies: TPS-a to TPS-h based on sequence properties and functional characteristics. Class I TPSs family contains , where class II contains TPS-c only which comprises genes for copalyl diphosphate related diterpene synthases. The TPS-d family mostly consists of bifunctional enzymes that are capable of the protonation-initiated cyclization, but also catalyze class I catalysis and mostly occur in gymnosperms. Whereas TPS-h is specific to the spikemoss Selaginella moellendorffii [10] . The TPS subfamily clades have been studied and recognized through sequencing and functional studies in a wide range of plants i.e., Arabidopsis thaliana containing ( 32 functional and 8 pseudo genes) [14], Vitis vinifera (69 putative functional and 63 pseudo genes) [15,16], Solanum lycopersicum (29 functional or potential functional genes and 15 mutated genes) [17], Oryza sativa (40 putative functional TPS genes) [18], Selaginella moellendorfii (18 TPS genes) [19], Cajanus cajan [20] and Populus trichocarpa (38 putative functional TPS genes) [21], Eucalyptus grandis (113 putative functional gene) Eucalyptus globulus (106 putative functional genes, 37 putative pseudogenes) [7]. Genome-wide analysis of terpene synthase genes family in different plants shows that all the TPS gene family is the mid-size with the number ranging Genome wide detection of O. sanctum terpene synthase genes PLOS ONE | wide detection of O. sanctum terpene synthase genes PLOS ONE |
doi:10.1371/journal.pone.0207097 fatcat:jjilzedjb5cdrn5ky67asplnv4