Juan-Juan Li1, Gan Zhang1, Ji-Hua Yu1, Yang-Yang Li1, Xin-He Huang1, Wan-Jun Wang1, Rui Tan1, Jia-Yu Zhou2, Hai Liao3. 1. College of Life Science and Engineering, Southwest Jiaotong University, Chengdu, 610031, Sichuan, China. 2. College of Life Science and Engineering, Southwest Jiaotong University, Chengdu, 610031, Sichuan, China. spinezhou@home.swjtu.edu.cn. 3. College of Life Science and Engineering, Southwest Jiaotong University, Chengdu, 610031, Sichuan, China. liaohai0918@163.com.
Abstract
OBJECTIVES: To clone and characterize caffeic acid 3-O-methyltransferase (LcCOMT) from the rhizome of Ligusticum chuanxiong, a traditional medicinal herb having a high content of ferulic acid. RESULTS: LcCOMT encoded an ORF of 362 amino acids with a calculated MW of 39,935 Da and pI of 5.94. Polygenetic tree indicated that LcCOMT was attributed to a new member of COMTs in plants. The recombinant LcCOMT was expressed in E. coli. HPLC and (1)H NMR analyses of purified LcCOMT protein confirmed that it could catalyze caffeic acid to produce ferulic acid in vitro. The further site-mutagenesis proved that His268 was one key catalytic residue. In addition, the substantial changing expression level of LcCOMT under chilling treatment suggested that LcCOMT might play important role in the accumulation of ferulic acid under chilling treatment. CONCLUSIONS: This is the first report of the isolation and characterization of a COMT clone from traditional medicine containing high contents of pharmaceutical ferulic acid.
OBJECTIVES: To clone and characterize caffeic acid 3-O-methyltransferase (LcCOMT) from the rhizome of Ligusticum chuanxiong, a traditional medicinal herb having a high content of ferulic acid. RESULTS: LcCOMT encoded an ORF of 362 amino acids with a calculated MW of 39,935 Da and pI of 5.94. Polygenetic tree indicated that LcCOMT was attributed to a new member of COMTs in plants. The recombinant LcCOMT was expressed in E. coli. HPLC and (1)H NMR analyses of purified LcCOMT protein confirmed that it could catalyze caffeic acid to produce ferulic acid in vitro. The further site-mutagenesis proved that His268 was one key catalytic residue. In addition, the substantial changing expression level of LcCOMT under chilling treatment suggested that LcCOMT might play important role in the accumulation of ferulic acid under chilling treatment. CONCLUSIONS: This is the first report of the isolation and characterization of a COMT clone from traditional medicine containing high contents of pharmaceutical ferulic acid.