title: Determination of D-aspartate N-methyltransferase activity in the starfish by direct analysis of N-methyl-D-aspartate with high-performance liquid chromatography.
abstract: We describe a method for the detection and quantification of D-aspartate N-methyltransferase activity. The enzyme catalyzes the S-adenosyl-L-methionine-dependent N-methylation of D-aspartate to form N-methyl-D-aspartate (NMDA). NMDA is detected directly by high-performance liquid chromatography (HPLC) of their (+)- and/or (-)-1-(9-fluorenyl)ethyl chloroformate fluorescent derivatives. The NMDA production in the assay mixture is linearly proportional to the incubation time and the amount of tissue homogenate. Using a 10 min incubation time, the method allows detection of the enzyme activity below 10 fmol/min. It can be used to analyze kinetic behavior and to quantify the enzyme from a wide variety of organisms.
authors.affiliation: Department of Chemistry and Biochemistry, Fukushima National College of Technology, 30 Nagao, Iwaki, Fukushima 970-8034, Japan. shibata@fukushima-nct.ac.jp
authors.forename: Kimihiko
authors.initials: K
authors.lastname: Shibata
authors.forename: Noriko
authors.initials: N
authors.lastname: Sugaya
authors.forename: Wakana
authors.initials: W
authors.lastname: Ono
authors.forename: Katsumasa
authors.initials: K
authors.lastname: Abe
authors.forename: Shouji
authors.initials: S
authors.lastname: Takahashi
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Kera
mesh_terms: D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D057075:Enzyme Assays / Q000379:methods*; D007700:Kinetics; D008780:Methyltransferases / Q000032:analysis / Q000378:metabolism*; D016202:N-Methylaspartate / Q000032:analysis* / Q000378:metabolism; D013215:Starfish / Q000201:enzymology* / Q000378:metabolism
chemical_list: D016202:N-Methylaspartate; D008780:Methyltransferases