title: Essential region for 3-N methylation in N-methyltransferases involved in caffeine biosynthesis.
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.
authors.affiliation: Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, Akita 010-0195, Japan. koumno@akita-pu.ac.jp
authors.forename: Kouichi
authors.initials: K
authors.lastname: Mizuno
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Kurosawa
authors.forename: Yuko
authors.initials: Y
authors.lastname: Yoshizawa
authors.forename: Misako
authors.initials: M
authors.lastname: Kato
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002110:Caffeine / Q000096:biosynthesis*; D031663:Clarkia / Q000201:enzymology / Q000235:genetics; D003069:Coffee / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers; D008745:Methylation; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010957:Plasmids; D016415:Sequence Alignment; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid
chemical_list: D003069:Coffee; D017931:DNA Primers; D002110:Caffeine; D008780:Methyltransferases