Characterization And Heterologous Expression Of BAHD Acyltransferases From Celastrus Angulatus Max

In plant secondary metabolism,enzymatic acylation is a widespread substitution reaction remarkably involved in the diversification of natural substances.Esters represent the largest and most diverse class of secondary metabolites.The postmodification processes of various metabolites such as aliphatic hydrocarbons,alkaloids,lignin,terpenoids and aromatic alcohols are all catalyzed by BAHD acyltransferase.Post-modification of esters is depend on BAHD acyltransferases,named after the first four biochemically characterized enzymes of the group,which use acyl-Co A thioesters as donor molecules.At present,esters and other chemicals are primarily derived from petroleum sources,which are nonrenewable.The increasing demand and fast depleting petroleum resources pushes the need to find sustainable and renewable platforms for their production.Therefore,looking at biological routes as an alternative to chemical synthesis has become an inevitable trend,in which the excavation of key enzymes is crucial.In this study,35 sequences of BAHD acyltransferases were screened from the transcriptome data,after that basic bioinformatics analysis was performed.Subsequently,Function of 10 acyltransferases(Ca AT1-10)and six acyltransferases from different sources were identified with different types of acyl donors and acyl acceptors.And then,the enzymatic studies on acyltransferases with acylation activity were performed in vitro before yeast expression in vivo.The experimental results show that the six enzymes(Ca AT1,Ca AT9,Ca AT10,ATF1,SAAT and Li AAT-3),possess catalytic activity on the substrate types including fatty alcohol,aromatic alcohol and monoterpene alcohol.Besides,the DFGWG conserved domain,one of the two highly conserved domains of BAHD acyltransferase,had little effect on the catalytic activity of some enzymes.The results of the enzymatic properties of the six acyltransferases indicate that Ca AT1,Ca AT9,and Ca AT10 are benzoyl-Co A preferred acyltransferases,while ATF1,SAAT,and Li AAT-3 prefer acetyl-Co A or isobutyryl-Co A.Finally,the results of expression of acyltransferase in yeast showed that SAAT and Li AAT-3 could be used as ideal phenylethyl acyltransferases,which can replace the previously reported ATF1 and can significantly increase the esters production.The discovery of thses BAHD acyltransferases by the studies on characterizition,enzymatic properties and yeast expression could provide a new insights into the evolution and function of these acyltransferases related to ester compounds.