Effect of periodate-oxidized ATP and other nucleotides on firefly luciferase

Abstract

Addition of periodate-oxidized ATP (oATP) to firefly luciferase-containing reaction mixtures enhanced light production when the reaction mixture contained > ˜8 μM ATP. The time course of light production was changed from a flash pattern to a constant light output during incubations of 8 μM) were used. There is little effect of oATP on firefly luciferase activity at low ATP concentrations (<2 μM) which gave steady production of light. ATP did not antagonize the inactivation of firefly luciferase by oATP. When the oATP was chemically reduced with sodium borohydride (giving orATP), there was no inactivation of firefly luciferase on incubation. When orATP was used in a short incubation the enhancement of light production and time course change were the same as those observed with oATP. The corresponding AMP and adenosine compounds (o and or) were slightly inhibitory to firefly luciferase activity. ADP was without effect but both oADP and orADP enhanced light production. Of these periodate-oxidized ADP, AMP, and adenosine derivatives only oADP inactivated firefly luciferase. The activating effect can be explained by a change in the conformation of the enzyme-product complex so that the product is released faster. In addition there is an inactivation of the enzyme by certain periodate-oxidized nucleotides during longer incubations. © 1994 Academic Press.