ADENOSINE 5-TRIPHOSPHATE MODULATION OF NITROBENZYLTHIOINOSINE BINDING-SITES IN PLASMA-MEMBRANES OF BOVINE CHROMAFFIN CELLS

Nitrobenzylthioinosine (NBTI) is a high affinity probe for facilitated diffusion nucleoside transporters. Kinetic analysis of the binding of [H-3]NBTI to plasma membranes of chromaffin cells was conducted in the presence or absence of adenosine 5'-triphosphale (ATP). Similar curvilinear plots with a Hill number of 1.32 were obtained in both conditions. ATP significantly increased the number of NBTI binding sites in these preparations showing B-max values of 1.62 +/- 0.20 pmol/mg protein for controls and 3.22 +/- 0.31 pmol/mg protein in the presence of ATP. However, the affinity constant (K-D) was not significantly modified. The non-metabolizable ATP analogue, 5'-adenylyl imidodiphosphate (AMP-PNP) and diadenosine tetraphosphate (Ap(4)A) can mimic the stimulatory ATP effect, but adenosine monophosphate (AMP) has no effect on the NBTI binding to plasma membranes. These results indicate a modulatory role for ATP, non-hydrolysis dependent, on nucleoside transport in chromaffin cells. Therefore, a nucleotide binding site on the nucleoside transporter similar to that described for glucose transporter could be suggested.