Electron capture dissociation as structural probe for noncovalent gas-phase protein assemblies.
Electron capture dissociation (ECD) of proteins in Fourier transform ion cyclotron resonance mass spectrometry usually leads to charge reduction and backbone-bond cleavage, thereby mostly retaining labile, intramolecular noncovalent interactions. In this report, we evaluate ECD of the 84-kDa noncovalent heptameric gp31 complex and compare this with sustained off-resonance irradiation collisionally activated dissociation (SORI-CAD) of the same protein. Unexpectedly, the 21+ charge state of the gp31 oligomer exhibits a main ECD pathway resulting in a hexamer and monomer, disrupting labile, intermolecular noncovalent bonds and leaving the backbone intact. Unexpectedly, the charge separation over the two products is highly proportional to molecular weight. This indicates that a major charge redistribution over the subunits of the complex does not take place during ECD, in contrast to the behavior observed when using SORI-CAD. We speculate that the ejected monomer retains more of its original structure in ECD, when compared to SORI-CAD. ECD of lower charge states of gp31 does not lead to dissociation of noncovalent bonds. We hypothesize that the initial gas-phase structure of the 21+ charge state is significantly different from the lower charge states. These structural differences result in the different reaction pathways when using ECD.
Geels
Rimco B J
RB
FOM Institute for Atomic and Molecular Physics, Kruislaan 407, 1098 SJ Amsterdam, The Netherlands.
van der Vies
Saskia M
SM
Heck
Albert J R
AJ
Heeren
Ron M A
RM
D018833:Chaperonins / Q000737:chemistry / Q000378:metabolism; D019281:Dimerization; D004583:Electrons; D006023:Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D013058:Mass Spectrometry; D011485:Protein Binding; D017550:Spectroscopy, Fourier Transform Infrared
D006023:Glycoproteins; D018833:Chaperonins