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Evaluating anti-CD32b F(ab) conformation using molecular dynamics and small-angle X-ray scattering

Evaluating anti-CD32b F(ab) conformation using molecular dynamics and small-angle X-ray scattering
Evaluating anti-CD32b F(ab) conformation using molecular dynamics and small-angle X-ray scattering
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.
0006-3495
289-299
Sutton, Emma, Jill
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Bradshaw, Richard
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Orr, Christian
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Frendéus, Björn
c960f7ce-006a-4ea9-8aac-f6f0304a746c
Larsson, Gunilla
67258081-13c2-40e5-b599-d6509f04fe7f
Teige, Ingrid
c6bf25bd-f182-46ab-9b8e-bd0159a4544b
Cragg, Mark
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Tews, Ivo
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Essex, Jonathan W.
1f409cfe-6ba4-42e2-a0ab-a931826314b5
Sutton, Emma, Jill
3d0922bd-d83b-4d82-b2de-3346b82217c4
Bradshaw, Richard
5e37ccd1-f8a8-4eec-a205-d68b57a877f3
Orr, Christian
f64259af-4120-481a-8e12-11344d005de0
Frendéus, Björn
c960f7ce-006a-4ea9-8aac-f6f0304a746c
Larsson, Gunilla
67258081-13c2-40e5-b599-d6509f04fe7f
Teige, Ingrid
c6bf25bd-f182-46ab-9b8e-bd0159a4544b
Cragg, Mark
ec97f80e-f3c8-49b7-a960-20dff648b78c
Tews, Ivo
9117fc5e-d01c-4f8d-a734-5b14d3eee8dd
Essex, Jonathan W.
1f409cfe-6ba4-42e2-a0ab-a931826314b5

Sutton, Emma, Jill, Bradshaw, Richard, Orr, Christian, Frendéus, Björn, Larsson, Gunilla, Teige, Ingrid, Cragg, Mark, Tews, Ivo and Essex, Jonathan W. (2018) Evaluating anti-CD32b F(ab) conformation using molecular dynamics and small-angle X-ray scattering. Biophysical Journal, 115 (2), 289-299. (doi:10.1016/j.bpj.2018.03.040).

Record type: Article

Abstract

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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Accepted/In Press date: 27 March 2018
e-pub ahead of print date: 17 July 2018

Identifiers

Local EPrints ID: 422678
URI: https://eprints.soton.ac.uk/id/eprint/422678
ISSN: 0006-3495
PURE UUID: 52738713-ad59-4821-98db-283031da88e7
ORCID for Richard Bradshaw: ORCID iD orcid.org/0000-0002-8652-4301
ORCID for Christian Orr: ORCID iD orcid.org/0000-0002-6137-8969
ORCID for Mark Cragg: ORCID iD orcid.org/0000-0003-2077-089X
ORCID for Ivo Tews: ORCID iD orcid.org/0000-0002-4704-1139
ORCID for Jonathan W. Essex: ORCID iD orcid.org/0000-0003-2639-2746

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Date deposited: 30 Jul 2018 16:30
Last modified: 14 Mar 2019 01:52

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Contributors

Author: Emma, Jill Sutton
Author: Richard Bradshaw ORCID iD
Author: Christian Orr ORCID iD
Author: Björn Frendéus
Author: Gunilla Larsson
Author: Ingrid Teige
Author: Mark Cragg ORCID iD
Author: Ivo Tews ORCID iD

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