The multiple sequence analysis showed that the
The multiple sequence analysis showed that the putative amino A 77636 hydrochloride sequence of HbFDP had 80–99% identity to the FDPs from H. brasiliensis, Lupinus and T. wilfordii, Malus×domestica, Centella asiatica, M. sativa, M.recutita, accession H. annuus, and A. thaliana. The present results are in agreement with recent findings reported by Sun et al. (2013); Su et al. (2015) and Zhao et al. (2015). The sequence alignment of the putative amino acid with other plant FDP proteins indicated the presence of highly conserved peptide domains (I–VII) that are essential for catalytic/functional activity in HbFDP (Szkopinska and Pochocka, 2005). The amino acid sequence predicted from this species shares an extensive conserved region with FDPs from different plant species. Furthermore, N-terminal sequence of HbFDP has no transit peptides for targeting into organelles. However, the predicted amino acid sequence from the reported cDNA contains two conserved regions with a high level identity to a sequence at the active catalytic site. Also the sequence contained the conserved motifs important to catalyze the successive 1′-4 condensation of the 5-carbon IPP to allylic substrates geranyl-, farnesyl-, or geranylgeranyl-diphosphate that has been suggested to be involved in transmembrane catalytic activity. Bioinformatic analysis showed that comparison of the deduced amino acid of Hevea and other plant FDPs has shown amino acid conservation at the two hydrophobic regions which correspond to the potential transmembrane domains. It is interesting to note that plant FDP catalytic site consists of a large central cavity formed by mostly antiparallel alpha helices with two aspartate-rich regions (DDXX(XX)D) located on opposite walls. These residues mediate binding of prenyl phosphates via bridging Mg ions, inducing proposed conformational changes that close the active site to solvent, protecting and stabilizing reactive carbocation intermediates. The conservation of the five characteristic domains found in all FPSs characterized so far suggests that we cloned a cDNA encoding an active FPS polypeptide. The Tyr residues, which determines the allylic product chain length, is located five amino acids upstream of the first Asp-rich domain as already described for other eukaryotic FPSs (Ohnuma et al., 1996). Similar results were also reported recently (Sun et al., 2013, Su et al., 2015, Zhao et al., 2015). Most recently, it has been demonstrated that the highly conserved aspartate-rich motif in region II with the sequence DDXX(XX)D is known as FARM (first Asp-rich motif), which is highly conserved in all known prenyltransferases that has been designated as the chain length determination region (Sun et al., 2013, Ferriols et al., 2015, Su et al., 2015, Zhao et al., 2015). Further, the sequence DDXXD is also called SARM (second Asp-rich motif) which are characteristic of prenyltransferases that can be used to the synthesis of isoprenoid diphosphates (Ohnuma et al., 1996). A phylogenetic tree of FDP from different plant species was constructed to examine the evolutionary relationships. All of the plant farnesyl diphosphate synthase sequences were separated into four main clusters. Hevea FDP was found to be highly homologous (88% nucleotide sequence identity) to Lupinus FDP gene followed by other species including Arabidopsis. HbFDP clustered together with Fabaceae and Asteraceae plants while Arabidopsis clustered as single group in the tree. Notably, plants from each family had formed groups as a single cluster in the tree. These results suggest that all these plants had very close evolutionary relationships. Moreover, they belong to Angiospermae–Dicotyledon plant kingdom groups. The present results are in agreement with recent reports (Sun et al., 2013, Ferriols et al., 2015, Su et al., 2015, Zhao et al., 2015). Southern results suggested that in Hevea, FDP is encoded by a small gene family consisting of two members. As evidenced in Southern blot analysis, it is not surprising to detect more than one isoforms of FDP in plants as it has been previously suggested that subcellular compartmentation of different isoforms of the enzyme occurs (Cunillera et al., 1996). Little information is available concerning FPS plant gene family complexity. There is evidence that FDP is encoded by at least two distinct genes in guayule rubber (Pan et al., 1996) and Arabidopsis (Cunillera et al., 1996), rice (Sanmiya et al., 1999) and tomato (Gaffe et al., 2000). The diversity of isoprenoid compounds in plants suggests that these compounds occur in multi-branched isoprenoid pathways (Bach, 1987). It has long been known that FDP as well as other allylic diphosphates act as initiators of cis1′-4 polyprenyl condensations that are the hallmark of rubber biosynthesis (Pan et al., 1996). The pathway for rubber biosynthesis in H. brasiliensis is different from the pathway(s) leading to biosynthesis of other isoprenoid compounds (Suwanmanee et al., 2002). Recently, it has been demonstrated that A.thaliana encodes two farnesyl diphosphate synthase genes, FPS1 and FPS2 that contains isozyme FPS1L (mitochondrial), FPS1S and FPS2 (both cytosolic) (Closa et al., 2010, Keim et al., 2012). It is noteworthy to mention that HbFDP gene has a high similarity with Arabidopsis FPS2 gene. Most recently, Zhao et al. (2015) reported a small FPS gene family of T. wilfordii that encodes at least two genes (TwFPS1 and TwFPS2) and these two genes shared a high level of sequence homology in the coding region but not in noncoding regions. Similarly, Sun et al. (2013) reported three genes (MsFPPS, MsGPPS and MsGGPPS) encoding isoprenyl diphosphate synthases (IDS) from alfalfa via a PCR based approach.