龙眼Dlc4h基因组织特异性表达及其生物信息学分析

Tissue-specific Expression and Bioinformatics Analysis of Dlc4h in Dimocarpus longan Lour.

肉桂酸4-羟化酶(cinnamate-4-hydroxylase,c4h)是合成黄酮类化合物的关键酶之一。为了研究Dlc4h基因的结构及其在龙眼次级代谢产物合成中的功能,本研究采用实时荧光定量PCR (qRT-PCR)技术分析Dlc4h基因在龙眼根、茎、叶组织中的特异性表达,并对该基因进行了生物信息学分析。定量分析结果表明:Dlc4h基因在各组织中均有表达,表达量由高到低依次为根、茎、叶。生物信息学分析结果表明:该基因全长1 611 bp,编码536个氨基酸;蛋白质相对分子质量为61 389.45 Da,等电点(pI)为8.82;它是一种不含信号肽、有跨膜结构、定位于质膜的可溶性亲水蛋白;二级结构主要包括α-螺旋和无规则卷曲,并具有卷曲螺旋结构;采用SWISS-MODEL构建Dlc4h三级结构模型,运用Ramachandran评估表明该模型准确可靠;Dlc4h预测得到的配体结合位点分别为128Arg、145Val、146Phe、153Trp、157Arg、213Met、329Ile、332Ala、333Ala、336Thr、337Thr、340Ser、402His、467Pro、468Phe、473Arg、475Cys和476Pro。本研究的结果为进一步研究Dlc4h基因在龙眼黄酮类化合物生物合成途径中行使的功能提供依据。

Cinnamate-4-hydroxylase(c4h)is one of the key enzymes functioning in the synthesis of flavonoids.In order to study the structure of Dlc4h gene and its function in the synthesis of secondary metabolites in Dimocarpus longan Lour,the expression patterns of Dlc4h gene in root,stem and leaf tissues were analyzed by the quantitative real time PCR(qRT-PCR).And the bioinformatics analysis was also carried out to further investigate Dlc4h gene in this study.The Dlc4h gene was expressed in all tissues,with the highest expression level in root,followed by stem and leaf.The full length cDNA of the Dlc4h was 1611 bp,encoding the protein of 536 amino acids.The Dlc4h had a calculated molecular weight of 61389.45 Da,and the theoretical isoelectric point was 8.82.Dlc4h was a signal free peptide with transmembrane structure and it was a soluble hydrophilic protein located on the plasma membrane.The secondary structure was mainly composed of alpha helix and random curl.Ramachandran evaluation showed that the tertiary structure model constructed by SWISS-MODEL was reliable and its ligand binding sites were 128Arg,145Val,146Phe,153Trp,157Arg,213Met,329Ile,332Ala,333Ala,336Thr,337Thr,340Ser,402His,467Pro,468Phe,473Arg,475Cys and 476Pro.The results of this study provide a basis for further study of the function of Dlc4h gene in longan flavonoids respectively biosynthesis pathway.