Mammalian complex I (NADH:ubiquinone oxidoreductase) exists as a mixture of interconvertible active (A) and de-activated (D) forms. The A-form is capable of NADH:quinone-reductase catalysis, but not the D-form. Complex I from the bacterium Paracoccus denitrificans, by contrast, exists only in the A-form. This bacterial complex contains 32 fewer subunits than the mammalian complex. The question arises therefore if the structural complexity of complex I from higher organisms correlates with its ability to undergo the A/D transition. In the present study, it was found that complex I from the bacterium Escherichia coli and from non-vertebrate organisms (earthworm, lobster, and cricket) did not show the A/D transitions. Vertebrate organisms (carp, frog, chicken), however, underwent similar A/D transitions to those of the well-characterized bovine complex I. Further studies showed that complex I from the lower eukaryotes, Neurospora crassa and Yarrowia lipolytica, exhibited very distinct A/D transitions with much lower activation barriers compared to the bovine enzyme. The A/D transitions of complex I as they relate to structure and regulation of enzymatic activity are discussed.