The metal binding properties of TpTroA have been studied in detail [30, 31] . One puzzling observation that was unclear until now is whether or not Zn and Mn bind to the same site in TpTroA. Nuclear spin relaxation properties of resonances from protons close to the ion-binding sites can be selectively perturbed in the presence of paramagnetic ions [50] . The unpaired electrons of Mn 2+ can interact with protons close to Mn 2+ and the effect of the strong paramagnetic interaction of the Mn cation leads to specific changes in the corresponding NMR signals, which helps to locate the Mn 2+ binding site [51] . Our NMR data demonstrated that Zn 2+ -SsTroA and Mn 2+ -SsTroA have very similar tertiary structures. Introduction of Mn induced the disappearance of some cross-peaks and the loss of intensity of others. Moreover, the binding of metal ions induces chemical shift changes for some of the cross-peaks (highlighted in circles, Fig. 6 ). However, the Zn 2+ -SsTroA and Mn 2+ -SsTroA spectra are nearly identical (except for the cross-peaks that disappear from spectrum), indicating that Zn 2+ and Mn 2+ share the adjacent binding region or even the same site in SsTroA.