RS-8359

Selenium deficiency has been reported to result in an extraordinary decrease of glutathione peroxidase (GSH-Px) and, reversely, an increase of detoxifying enzymes such as glutathione-S-transferase (GST), uridine-5'-diphosphate glucuronosyltransferase (UGT), nicotinamide-dependent quinine oxidoreductase (NQO1; DT-diaphorase), and epoxide hydrolase without significantly affecting cytochrome P450 activity. However, little is known about the effects on aldehyde oxidase 1 (AOX1) activity towards various kinds of aldehydes and N-heterocyclic aromatic compounds. The aim of this study is to clarify the effects of selenium deficiency on AOX1 in rats. As expected, selenium deficiency was confirmed by the extremely low activity of GSH-Px along with the increased activities of GST and DT-diaphorase. AOX1 activity towards vanillin and (S)-RS-8359 was increased by selenium deficiency, and that corresponded to an increase of AOX1 protein level but not to a decreased AOX1 mRNA level. It has been documented that the assembly of the catalytically active holoenzyme forms of the molybdo-flavoenzyme family is very complex and is controlled through transcriptional and translational events by many gene products. In addition, selenium deficiency has been known to cause oxidative stress that leads to an increase of AOX1 activity. Furthermore, aldehyde oxidase homolog 1 (AOH1) with properties similar to AOX1 is present in rodent liver. All the reports suggest that the mechanisms by which selenium deficiency increases AOX1 activity are highly complicated and investigated from different points of view.

Aldehyde oxidase (AO) is a homodimer with a subunit molecular mass of approximately 150 kDa. Each subunit consists of about 20 kDa 2Fe-2S cluster domain storing reducing equivalents, about 40 kDa flavine adenine dinucleotide (FAD) domain and about 85 kDa molybdenum cofactor (MoCo) domain containing a substrate binding site. In order to clarify the properties of each domain, especially substrate binding domain, chimeric cDNAs were constructed by mutual exchange of 2Fe-2S/FAD and MoCo domains between monkey and rat. Chimeric monkey/rat AO was referred to one with monkey type 2Fe-2S/FAD domains and a rat type MoCo domain. Rat/monkey AO was vice versa. AO-catalyzed 2-oxidation activities of (S)-RS-8359 were measured using the expressed enzyme in Escherichia coli. Substrate inhibition was seen in rat AO and chimeric monkey/rat AO, but not in monkey AO and chimeric rat/monkey AO, suggesting that the phenomenon might be dependent on the natures of MoCo domain of rat. A biphasic Eadie-Hofstee profile was observed in monkey AO and chimeric rat/monkey AO, but not rat AO and chimeric monkey/rat AO, indicating that the biphasic profile might be related to the properties of MoCo domain of monkey. Two-fold greater V(max) values were observed in monkey AO than in chimeric rat/monkey AO, and in chimeric monkey/rat AO than in rat AO, suggesting that monkey has the more effective electron transfer system than rat. Thus, the use of chimeric enzymes revealed that 2Fe-2S/FAD and MoCo domains affect the velocity and the quantitative profiles of AO-catalyzed (S)-RS-8359 2-oxidation, respectively.