A nonisotopic assay for acetylserotonin methyltransferase

✍ Scribed by Osamu Suzuki; Kunio Yagi

• Publisher: Elsevier Science
• Year: 1978
• Tongue: English
• Weight: 333 KB
• Volume: 88
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(78)90459-1

✦ Synopsis

A nonisotopic assay for acetylserotonin methyltransferase (ASMT) was devised. Melatonin, the product of the enzyme reaction, is measured fluorometrically after its reaction with o-phthaldialdehyde (OPT). The reaction of melatonin with OPT is carried out in 1 N HCI to suppress the reaction of N-acetylserotonin, the substrate of ASMT, with OPT. The mixture is gassed with nitrogen just before incubation at 60°C for 60 min in order to secure the linear relationship between the concentration of melatonin and the fluorescence intensity. This method is much simpler than the isotopic assay and also has as much high sensitivity. Moreover. in this assay the enzyme can be well saturated with S-adenosylmethionine, whereas in the isotopic assay it cannot.

Acetylserotonin methyltransferase (EC 2.1.1.4) (ASMT), which exists in the pineal bodies and the retinas of many species of animals, is an enzyme catalyzing the formation of melatonin from N-acetylserotonin (1). Extensive works have been carried out on the circadian rhythm of this enzyme (2). For measurement of the enzyme activity, the isotopic assay with radioactive S-adenosylmethionine (AMe) has been exclusively used (1). However, we tried to devise a nonisotopic method, and a sensitive and simple fluorometric assay for ASMT is presented in this paper.

Methods

Melatonin, o-phthaldialdehyde (OPT), cysteine, and 2-mercaptoethanol were of reagent grade (Nakarai Chemicals, Ltd.