A Novel Class of Plant Type III Polyketide Synthase Involved in Orsellinic Acid Biosynthesis from Rhododendron dauricum

Abstract

Rhododendron dauricum L. produces daurichromenic acid, the anti-HIV meroterpenoid consisting of sesquiterpene and orsellinic acid (OSA) moieties. To characterize the enzyme responsible for OSA biosynthesis, a cDNA encoding a novel polyketide synthase (PKS), orcinol synthase (ORS), was cloned from young leaves of R. dauricum. The primary structure of ORS shared relatively low identities to those of PKSs from other plants, and the active site of ORS had a unique amino acid composition. The bacterially expressed, recombinant ORS accepted acetyl-CoA as the preferable starter substrate, and produced orcinol as the major reaction product, along with four minor products including OSA. The ORS identified in this study is the first plant PKS that generates acetate-derived aromatic tetraketides, such as orcinol and OSA. Interestingly, OSA production was clearly enhanced in the presence of Cannabis sativa olivetolic acid cyclase, suggesting that the ORS is involved in OSA biosynthesis together with an unidentified cyclase in R. dauricum.

Complex Sexual Deception in an Orchid Is Achieved by Co-opting Two Independent Biosynthetic Pathways for Pollinator Attraction

“…We have observed that, among the transcripts showing strong differential expression in the C. plicata clubs tissue, there is a putative PKS gene. Plant PKSs are known to catalyze the formation of a plethora of natural products, including plant volatiles [63], and recent studies have now shown that acetophenones, such as phloroacetophenone (2,4,6-trihydroxyacetophenone), are novel catalytic products of plant PKS [64] (Table 1). Alternatively, biosynthesis of acetophenones may involve coenzyme A (CoA)-dependent b-oxidation of phenylpropanoid pathway precursors, as has been highlighted in plants such as tobacco [65] and white spruce [66].…”

Identification and Characterization of Daurichromenic Acid Synthase Active in Anti-HIV Biosynthesis

“…The FPKM value for RdFADOX1 was 74.7, which was much higher than those calculated for RdFADOX2 and RdFADOX3 (11.2 and 29.6,respectively). Notably, the gene for orcinol synthase (Taura et al, 2016; GenBank accession no. LC133082), the polyketide synthase involved in orsellinic acid biosynthesis in the DCA pathway, also is a highly expressed gene with an FPKM value of 81.9.…”

“…The gene-specific oligonucleotide primers and PCR conditions listed in Supplemental Table S3 were used for amplification of the RdFADOX1 to RdFADOX3 gene fragments. To obtain complete cDNA sequences, including untranslated regions, 3ʹ-and 5ʹ-RACE reactions were conducted as described previously (Taura et al, 2016). The cDNA template was prepared by RT of young leaf RNA using ReverTra Ace (Toyobo) and dT 17 AP primer.…”

“…The column and mobile phase were the same as those used for the HPLC analysis. The high-resolution MS (negative), MS/ MS, and UV spectra were collected as described previously (Taura et al, 2016).…”

How structural subtleties lead to molecular diversity for the type III polyketide synthases

“…are produced by type III PKS 3 -superfamily enzymes by iterative condensations of starter and extender acyl-CoA thioesters. Thus, in contrast to bacterial modular type I and dissociated type II PKS megaenzyme systems with multiple catalytic domains, such as ketoacyl synthase, acyltransferase, and acyl carrier protein (1,2), the structurally simple type III PKSs are homodimeric proteins of ϳ40-kDa subunits, which play a pivotal role in the remarkable structural diversification of polyketide backbone assemblies (3)(4)(5)(6)(7)(8)(9)(10)(11)(12). Each independent ketoacyl synthase domain in the type III PKS commonly maintains a "Cys-His-Asn" catalytic triad to catalyze the tethering of a CoA-linked starter substrate to the active-site Cys, which is followed by carbon-carbon (C-C) bond formation via decarboxylative Claisen-type condensation with an extender substrate (normally malonyl-CoA).…”