中药材2024，Vol.47Issue(11)：2680-2688,9.DOI:10.13863/j.issn1001-4454.2024.11.002
紫皮石斛DdPALs基因的克隆、表达及酶催化性质研究
Cloning,Expression and Enzyme-Catalyzed Properties of Dendrobium devonianum DdPALs Gene
摘要
Abstract
Objective:To explore the sequence structure and protein function of the PAL gene in Dendrobium devonianum,so as to provide a certain reference for the subsequent in-depth studies on the function of PAL in Dendrobium devonianum.Methods:Dendrobium devonianum was used as the material for PAL gene screening based on the transcriptome data,the PAL gene coding region(ORF)was cloned and sequenced,and the protein induction and in vitro enzymatic reaction were verified.Results:The ORF region of three PAL genes(named DdPAL1,DdPAL2,DdPAL3)was cloned,with a length of 1 674 bp,1 674 bp,2 130 bp,encoding 557,557,709 amino acids,respectively.The instability coefficients were 33.55,34.04,32.46,respectively,fat index was 96.23,98.10,92.69,respectively.The three DdPALs proteins all had phenylalanine ammonia-lyase superfamily conserved domains,which were subcellular in cytoplasm,and had no transmembrane domains or signaling peptides.Through prokaryotic expression,only the supernatant proteins of DdPAL1 and DdPAL3 were obtained.In vitro enzymatic reactions showed that both DdPAL1 and DdPAL3 proteins could effectively catalyze the gen-eration of trans-cinnamic acid from L-phenylalanine,and DdPAL3 could also catalyze the generation of p-coumaric acid from L-tyrosine.Conclusion:In this study,the phenylalanine ammonia-lyase(PAL)gene in Dendrobium devonianum is cloned,and the sequence charac-teristics,physicochemical properties,expression pattern and protein catalytic function of DdPALs gene are analyzed,which provides a reference for further study on the regulation of DdPALs gene in the synthesis of lignin,flavonoids and other secondary metabolites of Dendrobium devonianum.关键词
紫皮石斛/苯丙氨酸解氨酶/原核表达/酶催化/生物信息学Key words
Dendrobium devonianum Paxt./Phenylalanine ammonia-lyase/Prokaryotic expression/Enzyme catalysis/Bioinformatics分类
中医学引用本文复制引用
金芯羽,韩庆莉,何霞红,施蕊,唐军荣..紫皮石斛DdPALs基因的克隆、表达及酶催化性质研究[J].中药材,2024,47(11):2680-2688,9.基金项目
国家现代农业产业技术体系资助项目(CARS-21) （CARS-21）
农民院士科技服务站资助项目(202301) （202301）
