title: The first committed step reaction of caffeine biosynthesis: 7-methylxanthosine synthase is closely homologous to caffeine synthases in coffee (Coffea arabica L.).
abstract: In coffee and tea plants, caffeine is synthesized from xanthosine via a pathway that has three methylation steps. We identified and characterized the gene encoding the enzyme for the first methylation step of caffeine biosynthesis. The full-length cDNA of coffee tentative caffeine synthase 1, CtCS1, previously isolated by the rapid amplification of cDNA ends was translated with an Escherichia coli expression system and the resultant recombinant protein was purified using Ni-NTA column. The protein renamed CmXRS1 has 7-methylxanthine synthase (xanthosine:S-adenosyl-L-methionine methyltransferase) activity. CmXRS1 was specific for xanthosine and xanthosine 5'-monophosphate (XMP) could not be used as a substrate. The K(m) value for xanthosine was 73.7 microM. CmXRS1 is homologous to coffee genes encoding enzymes for the second and third methylation steps of caffeine biosynthesis.
authors.affiliation: Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, 010-0195, Akita, Japan. koumno@akita-pu.ac.jp <koumno@akita-pu.ac.jp>
authors.forename: Kouichi
authors.initials: K
authors.lastname: Mizuno
authors.forename: Misako
authors.initials: M
authors.lastname: Kato
authors.forename: Fumi
authors.initials: F
authors.lastname: Irino
authors.forename: Naho
authors.initials: N
authors.lastname: Yoneyama
authors.forename: Tatsuhito
authors.initials: T
authors.lastname: Fujimura
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Ashihara
mesh_terms: D002110:Caffeine / Q000096:biosynthesis*; D003069:Coffee / Q000201:enzymology*; D007700:Kinetics; D008780:Methyltransferases / Q000378:metabolism*; D008969:Molecular Sequence Data; D010802:Phylogeny; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D003069:Coffee; D011994:Recombinant Proteins; D002110:Caffeine; C477002:7-methylxanthosine synthase; D008780:Methyltransferases; C120479:caffeine synthase