Construction of expression system of rabbit aldehyde oxidase cDNA for the clarification of species differences

Abstract

A remarkably large species difference in cinchonidine oxidation activity catalyzed by aldehyde oxidase (AO) has been known, in particular between rabbit and monkey. As the first step in clarifying the phenomenon from the view point of structures of the active site, we attempted to construct an expression system of rabbit AO cDNA. The nucleotide sequences of cloned full-length rabbit AO cDNA were determined and confirmed to agree completely with those of genome DNA. The expression system in Escherichia coli was constructed in reference to the previously established method for monkey AO. Both expressed rabbit and monkey AO proteins correctly reproduced the remarkable species differences observed in their liver cytosols towards cinchonidine and methotrexate. Namely, the expressed rabbit AO protein showed extremely high activities than did that of monkey AO. A difference in the structure of the active site might be responsible for the substrate-dependent species difference towards the relatively bulky molecules of cinchonidine and methotrexate. The use of molecular biology techniques will be very useful to verify the hypothesis.

Identification and Characterization of Aldehyde Oxidase 5 in the Pheromone Gland of the Silkworm (Lepidoptera: Bombycidae)

“…However, the other functions of these active AOXs have in insects is unknown. In particular, it is important to understand the structure–activity relationships of AOX binding and metabolism ( Beedham et al 1987 , Dalvie et al 2012 , Coelho et al 2015 ), species differences ( Beedham et al 1987 , Terao et al 2006 , Dalvie et al 2013 ), AOX variability ( Terao et al 2006 , Adachi et al 2007 , Hutzler et al 2014 , Choughule et al 2015 ), and in vitro activity of AOX ( Stell et al 1989 , Alfaro et al 2009 , Liu et al 2009 ). Silkworm has many types of AOXs.…”