A Mass‐Spectrometry‐Based Modelling Workflow for Accurate Prediction of IgG Antibody Conformations in the Gas Phase

Abstract

Immunoglobulins are biomolecules involved in defence against foreign substances. Flexibility is key to their functional properties in relation to antigen binding and receptor interactions. We have developed an integrative strategy combining ion mobility mass spectrometry (IM‐MS) with molecular modelling to study the conformational dynamics of human IgG antibodies. Predictive models of all four human IgG subclasses were assembled and their dynamics sampled in the transition from extended to collapsed state during IM‐MS. Our data imply that this collapse of IgG antibodies is related to their intrinsic structural features, including Fab arm flexibility, collapse towards the Fc region, and the length of their hinge regions. The workflow presented here provides an accurate structural representation in good agreement with the observed collision cross section for these flexible IgG molecules. These results have implications for studying other nonglobular flexible proteins.

Enhanced neutralization potency of an identical HIV neutralizing antibody expressed as different isotypes is achieved through genetically distinct mechanisms

“…In addition, the sequence of the IgG3 hinge is very different to those of IgG1 and IgA1, which likely results in a distinct structural conformation leading to differential affinity of the IgG3 Fab to antigens 18 . The Fab–Fab distance in IgG3 antibodies is far greater than other Ig antibodies which may further influence binding affinity and neutralization potential 17 – 19 . In addition to the long IgG3 hinge length and increased Fab–Fab angles, this region also possesses O-linked glycosylation sites 20 .…”

Computational Strategies and Challenges for Using Native Ion Mobility Mass Spectrometry in Biophysics and Structural Biology

“…spheroids), or as density maps. Furthermore, it can be important to consider multiple alternative starting structures to ensure that the space is suitably explored 45 . This is pertinent for proteins or complexes that are particularly flexible or are characterised by intrinsically dynamic regions, and where maybe only one particularly stable or abundant structure has been characterized previously e.g.…”

Software Requirements for the Analysis and Interpretation of Native Ion Mobility Mass Spectrometry Data

“…Many of the data from native MS and IM-MS experiments can be highly complementary to data obtained by other structural biology methods, and can be exploited in molecular modelling scenarios. CCSs can be used to validate structures produced by, for example, homology modelling, coarse-grained, or molecular dynamics simulations 60 . MS and IM-MS data can also be fed directly into integrative modelling software programs to guide the refinement of the most likely conformation(s) of quaternary assemblies 61 .…”