Cloning and characterization of a novel O-methyltransferase from Flammulina velutipes that catalyzes methylation of pyrocatechol and pyrogallol structures in polyphenols. Issue 7 (3rd July 2015)

• Record Type: Journal Article
• Authors: Kirita, Masanobu; Tanaka, Yoshihisa; Tagashira, Motoyuki; Kanda, Tomomasa; Maeda-Yamamoto, Mari
• Is Part Of: Bioscience, biotechnology, and biochemistry. Volume 79:Issue 7(2015)
• Page Start: 1111
• Page End: 1118
• Publication Date: 2015-07-03
• Subjects: Flammulina velutipes -- O-methyltransferase -- O-methylated EGCGs -- O-methylated polyphenols; Biotechnology -- Periodicals; Biochemistry -- Periodicals; 660.6
• DOI: 10.1080/09168451.2015.1015955
• Languages: English
• ISSNs: 0916-8451

Abstract

Abstract: A novel O -methyltransferase gene was isolated from Flammulina velutipes . The isolated full-length cDNA was composed of a 690-nucleotide open reading frame encoding 230 amino acids. A database search revealed that the deduced amino acid sequence was similar to those of other O -methyltransferases; the highest identity was only 61.8% with Laccaria bicolor . The recombinant enzyme was expressed by Escherichia coli . BL21 (DE3) was assessed for its ability to methylate (−)-epigallocatechin-3- O -gallate (EGCG). LC–TOF–MS and NMR revealed that the enzyme produced five kinds of O -methylated EGCGs: (−)-epigallocatechin-3- O -(3- O -methyl)gallate, (−)-epigallocatechin-3- O -(4- O -methyl)gallate, (−)-epigallocatechin-3- O -(3, 4- O -dimethyl)gallate, (−)-epigallocatechin-3- O -(3, 5- O -dimethyl)gallate, and (−)-4′- O -methylepigallocatechin-3- O -(3, 5- O -dimethyl)gallate. The substrate specificity of the enzyme for 20 kinds of polyphenols was assessed using the crude recombinant enzyme of O -methyltransferase. This enzyme introduced methyl group(s) into polyphenols with pyrocatechol and pyrogallol structures. Graphical abstract: : Chemical structures that can and cannot be catalyzed by Fv-OMT. Pyrogallol and pyrocatechol were recognized, whereas resorcinol, phloroglucinol, and phenol were not.