The Effect Of RXR Motif On The Specificity Of Sesquiterpene Synthase Catalytic Product

Terpenoids are commonly found in the secondary metabolites of plants and microbes.With the discovery of more and more terpenoids,the study of terpene synthase is more deeper,which not only helps us to understand the synthesis mechanism of terpenoids in theory,but also understand the relationship between terpenoid synthase structure and function,besides,it can also help us to study the application of drugs,flavors and other aspects further.The different catalytic mechanism of terpene synthase is the key to reveal the number and structure diversity of terpenoids.The studies of conserved motif DDXXD,NSE / DTE and RXR are more important.In this paper,we focus on conserved motif RXR in the terpene synthase,and the effect of the conserved motif on the specificity of the catalytic products in three different sesquiterpene synthase catalytic mechanisms was discussed.The following works have done:1.To investigate the effect of RXR motif on the specificity of sesquiterpene cyclase products,we selected the single mutations of Artemisia annua Amorpha-4,11-diene synthase(ADS)R262 and R264,and joint mutations of R262 K and T296 A ?R262K and T296 V ?R262K and T296 I.We found that side-chain amino acids are needed when farnesyl diphosphate(FPP)ionization and pyrophosphate groups are released.But the cyclization reaction intermediate nerolidyl diphosphate((R,E)-NPP)ionization without R262 can be achieved.It is speculated that T296 A / V / I can interact with R262 K through other aminosoons or substrate carbon skeletons,and the results of this interaction are related to substrate conformation.Joint mutations can not act on(E,E)-FPP,but it can catalytic(R,E)-NPP?(S,E)-NPP respectively,and produce amorpha-4,7(11)-diene and(E)-?-farnesene respectively,nerolidol and ?-bisabolol both exist in these two products.When joint mutations catalytic(Z,E)-FPP,it can inhibit the capture of water by cationic intermediates in the R262 K cyclization reaction.R264 K has no effect on the product.2.In order to investigate the effect of RXR motif on the specificity of trans sesquiterpene cyclase products,we selected trans-cyclosporine tobacco tobacco 5-epi-aristolochene synthase(TEAS)R264,R266 single mutations,We found that TEAS catalyzed(Z,E)-FPP undergoes cis-isomerization,R264 K can increase the formation of terpene alcohols in the cis-synthesis pathway,this suggesting that R266 K disrupts the fragile interaction with substrates and other amino acids,the enzyme loss the activity.3.The first R in the sesquiterpene synthase RXR motif can assist in the ionization of FPP and introduce the cationic intermediates formed by the cyclization of water molecules to produce terpene alcohols.The second R mechanism is not clear.Moreover,RXR is specific for substrate conformations.