The characterization of the interactions of Zn 2+ ions with imidazole ligands is vital for understanding the function of a plethora of zinc enzymes at the molecular level. The infrared multiple photon dissociation (IRMPD) spectrum of mass selected Zn 2+(imidazole) 4 cations, [ZnIm 4] 2+, was obtained in the 670–1840 cm −1 fingerprint range by coupling the infrared free electron laser (IR-FEL) at the Centre Laser Infrarouge d’Orsay (CLIO) with a quadrupole ion trap mass spectrometer equipped with an electrospray ionization (ESI) source. The experimental efforts are complemented by quantum chemical calculations for [ZnIm n ] 2+ with n = 1–4 at the B3LYP level using basis sets ranging from cc-pVDZ to aug-cc-pVTZ. By comparison with calculated linear absorption spectra, the transitions observed in the IRMPD spectrum are assigned to vibrational modes of the imidazole ligands. In combination, the experimental data and the calculations provide detailed information about structure, metal–ligand bonding, charge distribution, and binding energy of the [ZnIm 4] 2+ complex in the gas phase. The superior abundance of the n = 4 complex of [ZnIm n ] 2+ in the mass spectra of the ESI source is indicative of the preferred coordination number CN = 4 for Zn 2+ interacting with imidazole ligands in both the gas and the liquid phase. Comparison of the IR spectra of [ZnIm n ] 2+ with that of bare Im reveals the impact of the strong Zn 2+–Im interaction on the electronic, geometric, and vibrational structure of the aromatic ligands upon sequential filling of the first coordination shell. Comparison with the structural and vibrational properties of the imidazole cation demonstrates that the metal-to-ligand charge transfer in [ZnIm n ] 2+ is dominated by σ donation, whereas contributions from π donation are minor.