Suramin blocks nucleotide triphosphate binding to ribosomal protein L3 from Trypanoplasma borreli

Ribosomal protein L3 (L3) has been demonstrated to participate in formation of the peptidyltransferase center and is essential for its catalytic activ ity. In the present study we show that L3 is able to bind nucleotide tripho sphates with high and specific affinity in vitro. L3 was serendipitously id entified by screening of a genomic phage library from a primitive kinetopla stid flagellate Trypanoplasma borreli with the ATPase domain of the topoiso merase II gene as a probe. The cloned gene was overexpressed and purified a s a his-tag fusion protein in E. coli. Radioligand binding experiments, usi ng [gamma-S-35]ATP, showed that L3 is able to bind ATP but also GTP and UTP with similar high affinity (IC50 50-100 nm), while it has no ATPase activi ty. Furthermore, we showed that L3 has more than 500-fold higher affinity f or nucleotide triphosphates compared to the corresponding nucleotide monoph osphates and diphosphates. Molecular genetic and biochemical analyses allow ed us to localize the NTP binding domain of L3 to the N-terminal 296 residu es. Suramin, a polysulfonated naphthylamine derivative of urea, known for i ts chemotherapeutic effects completely inhibited the binding of [gamma-S-35 ]ATP at subclinical levels. Results obtained with surface plasmon resonance technology showed that suramin both forms weak multimolecular complexes wi th L3 and binds strongly to L3 in nearly stoichiometric amounts.