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Updated: Jun 13, 2026

Caffeine Extraction, Enzymatic Activity and Gene Expression of Caffeine Synthase from Plant Cell Suspensions
Published on: October 2, 2018
Essential region for 3-N methylation in N-methyltransferases involved in caffeine biosynthesis
Kouichi Mizuno1, Shin-ichi Kurosawa, Yuko Yoshizawa
1Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, Akita 010-0195, Japan. koumno@akita-pu.ac.jp
Abstract:
The caffeine biosynthetic pathway is composed of three methylation steps, and N-methyltransferase catalyzing each step has high substrate specificity. Since the amino acid sequences among coffee 7-methylxanthosine synthase (CmXRS1), theobromine synthase, and caffeine synthase are highly homologous to each other, these substrate specificities seem to be determined in a very restricted region. The analysis of site-directed mutants for CmXRS1 that naturally acts at the initial step, i.e., 7-N methylation of xanthosine, revealed that the activity of 3-N methylation needs a histidine residue at corresponding position 161 in the CmXRS1 sequence. We succeeded in producing the mutant enzyme which can catalyze the first and second methylation steps in caffeine biosynthesis.
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