Medical College of Wisconsin
CTSICores SearchResearch InformaticsREDCap

Dissecting the dynamic conformations of the metamorphic protein lymphotactin. J Phys Chem B 2014 Oct 30;118(43):12348-59

Date

09/27/2014

Pubmed ID

25259958

DOI

10.1021/jp504997k

Scopus ID

2-s2.0-84908577548   19 Citations

Abstract

A mass spectrometer provides an ideal laboratory to probe the structure and stability of isolated protein ions. Interrogation of each discrete mass/charge-separated species enables the determination of the intrinsic stability of a protein fold, gaining snapshots of unfolding pathways. In solution, the metamorphic protein lymphotactin (Ltn) exists in equilibrium between two distinct conformations, a monomeric (Ltn10) and a dimeric (Ltn40) fold. Here, we use electron capture dissociation (ECD) and drift tube ion mobility-mass spectrometry (DT IM-MS) to analyze both forms and use molecular dynamics (MD) to consider how the solution fold alters in a solvent-free environment. DT IM-MS reveals significant conformational flexibility for the monomer, while the dimer appears more conformationally restricted. These findings are supported by MD calculations, which reveal how salt bridges stabilize the conformers in vacuo. Following ECD experiments, a distinctive fragmentation pattern is obtained for both the monomer and dimer. Monomer fragmentation becomes more pronounced with increasing charge state especially in the disordered regions and C-terminal α-helix in the solution fold. Lower levels of fragmentation are seen in the β-sheet regions and in regions that contain salt bridges, identified by MD simulations. The lowest charge state of the dimer for which we obtain ECD data ([D+9H](9+)) exhibits extensive fragmentation with no relationship to the solution fold and has a smaller collision cross section (CCS) than charge states 10-13+, suggesting a "collapsed" encounter complex. Other charge states of the dimer, as for the monomer, are resistant to fragmentation in regions of β-sheets in the solution fold. This study provides evidence for preservation and loss of global fold and secondary structural elements, providing a tantalizing glimpse into the power of the emerging field of native top-down mass spectrometry.

Author List

Harvey SR, Porrini M, Konijnenberg A, Clarke DJ, Tyler RC, Langridge-Smith PR, MacPhee CE, Volkman BF, Barran PE

Author

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

Disulfides
Gases
Humans
Hydrogen Bonding
Lymphokines
Mass Spectrometry
Molecular Dynamics Simulation
Peptide Fragments
Protein Multimerization
Protein Stability
Protein Structure, Secondary
Sialoglycoproteins
jenkins-FCD Prod-444 eb4ebd1a08581aba961d3befd3b851a3c3ec6b46