Mint3 is known to enhance aerobic ATP production, known as the Warburg effect, by binding to FIH-1. Since this effect is considered to be beneficial for cancer cells, the interaction is a promising target for cancer therapy. However, previous research has suggested that the interacting region of Mint3 with FIH-1 is intrinsically disordered, which makes investigation of this interaction challenging. Therefore, we adopted a physicochemical approach that combined thermodynamic studies with structural analyses in solution, to clarify the binding mechanism. First, using a combination of CD, NMR, and hydrogen/deuterium exchange mass spectrometry (HDX-MS), we confirmed that the N-terminal half, which is the interacting part of Mint3, is mostly disordered. Next, using isothermal titration calorimetry (ITC), we revealed that the interaction of Mint3 and FIH-1 produces an enormous change in enthalpy and entropy. The profile is consistent with the model that the flexibility of disordered Mint3 is drastically reduced upon binding to FIH-1. Moreover, we performed a series of ITC experiments with several types of truncated Mint3s, an effective approach since the interacting part of Mint3 is disordered, and identified 78-88 as a novel core site for binding to FIH-1. The truncation study of Mint3 also revealed the thermodynamic contribution of each part to the interaction, where one contributes to the affinity (ΔG), while the other only affects enthalpy (ΔH), by forming non-covalent bonds. This insight can serve as a foothold for further investigation of IDRs and drug development for cancer therapy.
NEDD4: A Promising Target for Cancer Therapy
