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

Abstract

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-Ap 3A 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 (Ap 3N)-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 V max and K m 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-Ap 3A signaling complex lifetime leading to alteration of its antitumor activity.