Abstract
Acireductone dioxygenase (ARD) is an enzyme found in the highly-conserved methionine salvage pathway. ARD is the only known metalloenzyme to perform different biochemical activities depending on the identity of the bound metal ion. The native, or on-pathway, activity is catalyzed by ferrous (Fe2+ -bound) ARD, producing the ketoacid precursor of methionine, while ferric (Fe3+-bound) ARD has no activity. Non-native, or off-pathway, activity has been previously identified when ARD is bound to Ni2+, Co2+, Mn2+, and yields off-pathway products including formate, methylthiopropionate, and carbon monoxide, the latter of which has been implicated in cell signals leading to tumor progression. ARD has also been previously implicated in a number of other cellular roles, though it is unclear how all of these roles may be related. For the first reported time, iron-containing MmARD has been successfully reduced in situ rather than in an anaerobic chamber. This work provides further insight into the on-pathway form of ARD in the form of kinetic and thermal stability studies, as well as efforts towards obtaining a crystal structure of ferrous iron-containing ARD. Additionally, the inactive ferric iron-containing ARD has been crystallized and its thermal stability investigated.