Biomolecular recognition of vancomycin antibiotics with its cell-wall precursor analogue Ac(2)(L)K(D)A(D)A has been investigated in the gas phase through a combined laser spectroscopy/mass spectrometry approach. The mid-IR spectra (1100-1800 cm(-1)) of these mass-selected anionic species have been recorded by means of resonant infrared multiphoton dissociation (IRMPD) spectroscopy performed with the free-electron laser CLIO. Structural assignment has been achieved through comparisons with the low-energy conformers obtained from replica-exchange molecular dynamics simulations, for which IR spectra were calculated using a hybrid quantum mechanics/semi-empirical (QM/SE) method at the DFT/B3LYP/6-31+G*/AM1 level. Comparison between deprotonated vancomycin and its non-covalently bound V + Ac(2)(L)K(D)A(D)A complex shows significant spectral shifts of the carboxylate stretches and the Amide I and Amide II modes that are satisfactorily reproduced in the structures known from the condensed phase. Both theoretical and experimental findings provide strong evidence that the native structure of the deprotonated V + Ac(2)(L)K(D)A(D)A complex is preserved in the gas phase.