Ubc9-induced inhibition of diadenosine triphosphate hydrolase activity of the putative tumor suppressor protein Fhit

Fhit protein is the product of the putative tumor suppressor fragile histidine triad (FHIT) gene. The way by which Fhit exerts its antitumor activity remains largely unknown, although the Fhit-Ap3A complex is believed to be the native signaling form of Fhit. Here, we have shown that Fhit protein interacts with hUbc9, a recombinant human SUMO-1 conjugating enzyme, in an adenosine(5')triphospho(5')nucleoside (Ap3N)-dependent manner. Our experiments showed that the dinucleoside polyphosphate hydrolase activity of Fhit is suppressed by interacting with hUbc9 protein. In the presence of equimolar hUbc9 the Vmax and Km activity of Fhit was decreased by 35%. Analysis of Fhit kinetics in the presence of different fixed concentrations of Ubc9 showed that Ubc9 is an uncompetitive inhibitor. Including SUMO-1 protein in the assay neither affected the Fhit activity nor modified the effect of Ubc9 on Fhit kinetics. Our data suggest that hUbc9-induced inhibition of Fhit may result in an elongation of the Fhit-Ap3A signaling complex lifetime leading to alteration of its antitumor activity.