Medical College of Wisconsin
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Electron capture dissociation and drift tube ion mobility-mass spectrometry coupled with site directed mutations provide insights into the conformational diversity of a metamorphic protein. Phys Chem Chem Phys 2015 Apr 28;17(16):10538-50



Pubmed ID




Scopus ID

2-s2.0-84928027570   9 Citations


Ion mobility mass spectrometry can be combined with data from top-down sequencing to discern adopted conformations of proteins in the absence of solvent. This multi-technique approach has particular applicability for conformationally dynamic systems. Previously, we demonstrated the use of drift tube ion mobility-mass spectrometry (DT IM-MS) and electron capture dissociation (ECD) to study the metamorphic protein lymphotactin (Ltn). Ltn exists in equilibrium between distinct monomeric (Ltn10) and dimeric (Ltn40) folds, both of which can be preserved and probed in the gas-phase. Here, we further test this mass spectrometric framework, by examining two site directed mutants of Ltn, designed to stabilise either distinct fold in solution, in addition to a truncated form consisting of a minimum model of structure for Ltn10. The truncated mutant has similar collision cross sections to the wild type (WT), for low charge states, and is resistant to ECD fragmentation. The monomer mutant (CC3) presents in similar conformational families as observed previously for the WT Ltn monomer. As with the WT, the CC3 mutant is resistant to ECD fragmentation at low charge states. The dimer mutant W55D is found here to exist as both a monomer and dimer. As a monomer W55D exhibits similar behaviour to the WT, but as a dimer presents a much larger charge state and collision cross section range than the WT dimer, suggesting a smaller interaction interface. In addition, ECD on the W55D mutant yields greater fragmentation than for the WT, suggesting a less stable β-sheet core. The results highlight the power of MS to provide insight into dynamic proteins, providing further information on each distinct fold of Ltn. In addition we observe differences in the fold stability following single or double point mutations. This approach, therefore, has potential to be a useful tool to screen for the structural effects of mutagenesis, even when sample is limited.

Author List

Harvey SR, Porrini M, Tyler RC, MacPhee CE, Volkman BF, Barran PE


Brian F. Volkman PhD Professor in the Biochemistry department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Mass Spectrometry
Models, Molecular
Mutagenesis, Site-Directed
Protein Conformation
Protein Unfolding
jenkins-FCD Prod-461 7d7c6113fc1a2757d2947d29fae5861c878125ab