Human thymidylate synthase is a homodimeric enzyme that plays a key role in DNA synthesis and is a target for several clinically important anticancer drugs that bind to its active site. We have designed peptides to specifically target its dimer interface. Here we show through X-ray diffraction, spectroscopic, kinetic, and calorimetric evidence that the peptides do indeed bind at the interface of the dimeric protein and stabilize its di-inactive form. The "LR" peptide binds at a previously unknown binding site and shows a previously undescribed mechanism for the allosteric inhibition of a homodimeric enzyme. It inhibits the intracellular enzyme in ovarian cancer cells and reduces cellular growth at low micromolar concentrations in both cisplatin-sensitive and -resistant cells without causing protein overexpression. This peptide demonstrates the potential of allosteric inhibition of hTS for overcoming platinum drug resistance in ovarian cancer.
Crystal Structure of histidine-tagged human thymidylate synthase / Pozzi, Cristina; Cardinale, Daniela; Guaitoli, G.; Tondi, Donatella; Luciani, Rosaria; Myllykallio, H.; Ferrari, Stefania; Costi, Maria Paola; Mangani, S.. - (2011), pp. 1-1. [10.2210/pdb3n5g/pdb]