Identification of the missing component in the mitochondrial benzamidoxime prodrug-converting system as a novel molybdenum enzyme.
Amidoximes can be used as prodrugs for amidines and related functional groups to enhance their intestinal absorption. These prodrugs are reduced to their active amidines. Other N-hydroxylated structures are mutagenic or responsible for toxic effects of drugs and are detoxified by reduction. In this study, a N-reductive enzyme system of pig liver mitochondria using benzamidoxime as a model substrate was identified. A protein fraction free from cytochrome b5 and cytochrome b5 reductase was purified, enhancing 250-fold the minor benzamidoxime-reductase activity catalyzed by the membrane-bound cytochrome b5/NADH cytochrome b5 reductase system. This fraction contained a 35-kDa protein with homologies to the C-terminal domain of the human molybdenum cofactor sulfurase. Here it was demonstrated that this 35-kDa protein contains molybdenum cofactor and forms the hitherto ill defined third component of the N-reductive complex in the outer mitochondrial membrane. Thus, the 35-kDa protein represents a novel group of molybdenum proteins in eukaryotes as it forms the catalytic part of a three-component enzyme complex consisting of separate proteins. Supporting these findings, recombinant C-terminal domain of the human molybdenum cofactor sulfurase exhibited N-reductive activity in vitro, which was strictly dependent on molybdenum cofactor.
Havemeyer
Antje
A
Pharmaceutical Institute, Department of Pharmaceutical and Medicinal Chemistry, Christian-Albrechts-University of Kiel, 24118 Kiel, Germany.
Bittner
Florian
F
Wollers
Silke
S
Mendel
Ralf
R
Kunze
Thomas
T
Clement
Bernd
B
D000818:Animals; D001550:Benzamidines / Q000378:metabolism*; D003067:Coenzymes / Q000378:metabolism; D008930:Mitochondria, Liver / Q000201:enzymology*; D015394:Molecular Structure; D008982:Molybdenum / Q000378:metabolism*; D011355:Prodrugs / Q000378:metabolism*; D013466:Sulfurtransferases / Q000378:metabolism*; D013552:Swine
D001550:Benzamidines; D003067:Coenzymes; D011355:Prodrugs; C053561:benzamidoxime; D008982:Molybdenum; D013466:Sulfurtransferases