Evaluating Anti-CD32b F(ab) Conformation Using Molecular Dynamics and Small-Angle X-Ray Scattering.

Authors:
Emma J Sutton
Emma J Sutton
North Staffordshire CCG and Stoke-on-Trent CCG
Richard T Bradshaw
Richard T Bradshaw
School of Chemistry
Lawrence | United States
Christian M Orr
Christian M Orr
University of Southampton Faculty of Medicine
United Kingdom
Gunilla Larsson
Gunilla Larsson
Swedish National Rett Centre
Singapore | Singapore
Ingrid Teige
Ingrid Teige
Research
Dallas | United States
Mark S Cragg
Mark S Cragg
University of Southampton
Highfield | United Kingdom
Ivo Tews
Ivo Tews
Heidelberg University Biochemistry Center
Germany

Biophys J 2018 Jul;115(2):289-299

Department of Chemistry, University of Southampton, Highfield Campus, Southampton, United Kingdom. Electronic address:

Complementary strategies of small-angle x-ray scattering (SAXS) and crystallographic analysis are often used to determine atomistic three-dimensional models of macromolecules and their variability in solution. This combination of techniques is particularly valuable when applied to macromolecular complexes to detect changes within the individual binding partners. Here, we determine the x-ray crystallographic structure of a F(ab) fragment in complex with CD32b, the only inhibitory Fc-γ receptor in humans, and compare the structure of the F(ab) from the crystal complex to SAXS data for the F(ab) alone in solution. We investigate changes in F(ab) structure by predicting theoretical scattering profiles for atomistic structures extracted from molecular dynamics (MD) simulations of the F(ab) and assessing the agreement of these structures to our experimental SAXS data. Through principal component analysis, we are able to extract principal motions observed during the MD trajectory and evaluate the influence of these motions on the agreement of structures to the F(ab) SAXS data. Changes in the F(ab) elbow angle were found to be important to reach agreement with the experimental data; however, further discrepancies were apparent between our F(ab) structure from the crystal complex and SAXS data. By analyzing multiple MD structures observed in similar regions of the principal component analysis, we were able to pinpoint these discrepancies to a specific loop region in the F(ab) heavy chain. This method, therefore, not only allows determination of global changes but also allows identification of localized motions important for determining the agreement between atomistic structures and SAXS data. In this particular case, the findings allowed us to discount the hypothesis that structural changes were induced upon complex formation, a significant find informing the drug development process. The methodology described here is generally applicable to deconvolute global and local changes of macromolecular structures and is well suited to other systems.

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Evaluating Anti-CD32b F(ab) Conformation


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http://dx.doi.org/10.1016/j.bpj.2018.03.040DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6050753PMC

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July 2018
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