Characterization of the pH‐dependent dissociation of a multimeric metalloprotein Streptomyces rubiginosus xylose isomerase by ESI FT‐ICR mass spectrometry

We report an analysis of the pH‐dependent dissociation of a multimeric metalloprotein, xylose isomerase from Streptomyces rubiginosus (XI), by electrospray ionization (ESI) Fourier transform ion cyclotron resonance (FT‐ICR) mass spectrometry. Xylose isomerases are industrially significant enzymes that catalyze interconversion of aldose and ketose sugars. XI is biologically active as a ∼173‐kDa tetrameric complex, comprised of four identical ∼43‐kDa subunits and eight metal cations, unequivocally identified as the Mg2+ cations in this work. ESI FT‐ICR mass spectra of XI measured in the pH range of 3.0–6.9 indicated that the dissociation of the intact holo‐tetramer is initiated by the loss of all eight Mg2+ cations at pH ≤5.0, followed by step‐by‐step dissociation of the remaining apo‐tetramer to trimers, dimers and monomers. In addition, a ∼346‐kDa protein octamer was detected at pH 6.9.