Affiliation:
1. Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences Key Laboratory of Beijing for Identification and Safety Evaluation of Chinese Medicine, , No. 16, Nanxiaojie, Dongzhimennei, Beijing 100700, China
2. Amway (China) Botanical R&D Centre , Wuxi 214115, China
Abstract
Abstract
Lotus (Nelumbo nucifera), an ancient aquatic plant, possesses a unique pharmacological activity that is primarily contributed by benzylisoquinoline alkaloids (BIAs). However, only few genes and enzymes involved in BIA biosynthesis in N. nucifera have been isolated and characterized. In the present study we identified the regiopromiscuity of an O-methyltransferase, designated NnOMT6, isolated from N. nucifera; NnOMT6 was found to catalyze the methylation of monobenzylisoquinoline 6-O/7-O, aporphine skeleton 6-O, phenylpropanoid 3-O, and protoberberine 2-O. We further probed the key residues affecting NnOMT6 activity via molecular docking and molecular dynamics simulation. Verification using site-directed mutagenesis revealed that residues D316, N130, L135, N176A, D269, and E328 were critical for BIA O-methyltransferase activities; furthermore, N323A, a mutant of NnOMT6, demonstrated a substantial increase in catalytic efficiency for BIAs and a broader acceptor scope compared with wild-type NnOMT6. To the best of our knowledge, this is the first study to report the O-methyltransferase activity of an aporphine skeleton without benzyl moiety substitutions in N. nucifera. The study findings provide biocatalysts for the semisynthesis of related medical compounds and give insights into protein engineering to strengthen O-methyltransferase activity in plants.
Publisher
Oxford University Press (OUP)
Subject
Horticulture,Plant Science,Genetics,Biochemistry,Biotechnology
Cited by
4 articles.
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