Oxidosqualene Cyclases Involved in the Biosynthesis of Diverse Triterpenes in Camellia sasanqua

Abstract

Camellia sasanqua is an important economic plant that is rich in lipophilic triterpenols with pharmacological activities including antiallergic, anti-inflammatory, and anticancer activities. However, the key enzymes related to triterpene biosynthesis have seldom been studied in C. sasanqua. Oxidosqualene cyclases (OSCs) are the rate-limiting enzymes related to triterpene biosynthesis. In this study, seven putative OSC genes (CsOSC1-7) were mined from the C. sasanqua transcriptome. Six CsOSCs were characterized for the biosynthesis of diverse triterpene skeletons, including α-amyrin, β-amyrin, δ-amyrin, dammarenediol-II, ψ-taraxasterol, taraxasterol, and cycloartenol by the heterologous expression system. CsOSC3 was a multiple functional α-amyrin synthase. Three key residues, Trp260, Tyr262, and Phe415, are critical to the catalytic performance of CsOSC3 judging from the results of molecular docking and site-directed mutagenesis. These findings provide important insights into the biosynthesis pathway of triterpenes in C. sasanqua.

Genome-Wide Investigation of Oxidosqualene Cyclase Genes Deciphers the Genetic Basis of Triterpene Biosynthesis in Tea Plants

“…Likewise, CsOSC6 was identified as a multifunctional α-amyrin synthase, producing β-amyrin (1), α-amyrin (2), and an unknown lanostane derivative (4) at a ratio of 33:64:3 (Figures 2D, S3 and Table S4). CsOSC8 showed activity resembling a known cycloartenol synthase (UXE43252) from C. sasanqua 23 (Figures 2E and S4), in agreement with their phylogenetic grouping.…”

“…CsOSC7 and CsOSC8 are grouped alongside known cycloartenol synthases. It is interesting to note that CsOSCs showed a high degree of similarity with OSC proteins from C. sasanqua, 23 a phylogenetically related species to C. sinensis. Given the substantial evolutionary relatedness of OSC enzymes across plant lineages, a high degree of sequence conservation among these enzymes would be indicative of functional similarities.…”

“…All CsOSCs form extensive hydrophobic interactions between multiple conserved residues and the substrate backbone (Figure S6A−D). Within the signature MWCYCR motif of β-amyrin synthases, 16 key interactions were observed, 23 The numbered compounds are as follows: 1, camelliol C; 2, lupane derivative 1; 3, oleanane derivative; 4, δ-amyrin; 5, βamyrin; 6, lupane derivative 2; 7, α-amyrin; 8, lupeol; 9, ψ-taraxasterol; 10, lanostane derivative 1; 11, lanostane derivative 2; 12, lupane derivative 3; 13, friedelin; 14, taraxasterol; 15, 28,29-bis-nor-24-methylcycloart-3,6-diol-22-en isomer; 16, 28,29-bis-nor-24-methylcycloart-3,6-diol-22-en; 17, cycloartenol/cycloeucalenol, 18, roxburghiadiol B; and 19, 28,29-bis-nor-24-methylcycloart-3β,6α-diol. S7).…”

“…40 Mutations of the corresponding residues, such as W260L in a multifunctional αamyrin synthase from C. sasanqua and W257L in a β-amyrin synthase from Perilla frutescens, demonstrated similar effects. 23,41 The Y261H mutation in CsOSC1, also located within the MWCYCR motif, resulted in a rapid loss of the enzyme activity, where the β-amyrin production was completely abolished (Figures 6A,6E, S7 and Table S8). This is consistent with previous research that observed the loss of β-amyrin synthesis upon equivalent mutations of Tyr to aliphatic residues (Ala, Val, Leu, and Ile) or His in the β-amyrin synthase from Euphorbia tirucalli (EtAS).…”

Triterpenoids from the leaves of Camellia chrysantha growing in Quang Ninh (Vietnam) and their activities on main protease (Mpro) and ACE2

“…[ 19 ] revealed that this compound was β‐amyrin. It was detected in C. japonica stem bark; [ 17 ] C. sinensis leaf; [ 18,20 ] C. tenuifolia seed oil; [ 21 ] in the flowers and stem of C. sativa ; [ 22 ] in the roots, leaves and stem of C. sasanqua ; [ 23 ] in the fruits of C. chekiangoleosa , C. polyodonta , C. semiserrata and C. reticulata . [ 24 ] Furthermore, β‐amyrin was splited for the first time from C. chrysantha .…”