A Novel Assay Method to Determine the β-Elimination of Se-Methylselenocysteine to Monomethylselenol by Kynurenine Aminotransferase 1

• Main Authors: Arun Kumar Selvam (Author), Mikael Björnstedt (Author)
• Format: Book
• Published: MDPI AG, 2020-02-01T00:00:00Z.
• Subjects: article
• Online Access: Connect to this object online.
• Notes: 2076-3921; 10.3390/antiox9020139
• Keywords: kynurenine aminotransferase 1; se-methylselenocysteine; β-elimination activity; methylselenol; thioredoxin reductase; kyat1 inducers; and kyat1 inhibitors; Therapeutics. Pharmacology; RM1-950
• Source: Antioxidants, Vol 9, Iss 2, p 139 (2020)

Kynurenine aminotransferase 1 (KYAT1 or CCBL1) plays a major role in Se-methylselenocysteine (MSC) metabolism. It is a bi-functional enzyme that catalyzes transamination and beta-elimination activity with a single substrate. KYAT1 produces methylselenol (CH₃SeH) via β-elimination activities with MSC as a substrate. This methylated selenium compound is a major cytotoxic selenium metabolite, causing apoptosis in a wide variety of cancer cells. Methylselenol is volatile and possesses extraordinary nucleophilic properties. We herein describe a simple spectrophotometric assay by combining KYAT1 and thioredoxin reductase (TrxR) to detect CH₃SeH in a coupled activity assay. The metabolite methylselenol and its oxidized form from MSC metabolism is utilized as a substrate for TrxR1 and this can be monitored spectroscopically at 340 nm. Our results show the feasibility of monitoring the β-elimination of KYAT1 by our assay and the results were compared to the previously described β-elimination assays measuring pyruvate. By using known inhibitors of KYAT1 and TrxR1, we further validated the respective reaction. Our data provide a simple but accurate method to determine the β-elimination activity of KYAT1, which is of importance for mechanistic studies of a highly interesting selenium compound.