Comparison of the pH- and thermally-induced fluctuations of a therapeutic antibody Fab fragment by molecular dynamics simulation

Computational and Structural Biotechnology Journal

Nuria Codina¹  Nesrine Chakroun¹  Cheng Zhang¹  Paul A. Dalby¹  Haoran Yu²  Frank Kozielski³  Jiazhi Tang³ 

[1] Department of Biochemical Engineering, University College London, Gordon Street, London WC1E 7JE, United Kingdom;Department of Chemistry, University College London, 20 Gordon Street, London, WC1H 0AJ, United Kingdom;Department of Pharmaceutical and Biological Chemistry, School of Pharmacy, University College London, 29–39 Brunswick Square, London WC1N 1AX, United Kingdom;

关键词: Antibody fragment;    Crystal structure;    Protein stability;    Protein aggregation;    Protein engineering;    Molecular dynamics simulations;   

DOI  :  

来源: DOAJ

【 摘 要 】

Successful development of protein therapeutics depends critically on achieving stability under a range of conditions. A deeper understanding of the drivers of instability across different stress conditions, will enable the engineering of more robust protein scaffolds. We compared the impacts of low pH and high temperature stresses on the structure of a humanized antibody fragment (Fab) A33, using atomistic molecular dynamics simulations, using a recent 2.5 Å crystal structure. This revealed that low-pH induced the loss of native contacts in the domain CL. By contrast, thermal stress led to 5–7% loss of native contacts in all four domains, and simultaneous loss of >30% of native contacts in the VL-VH and CL-CH interfaces. This revealed divergent destabilising pathways under the two different stresses. The underlying cause of instability was probed using FoldX and Rosetta mutation analysis, and packing density calculations. These agreed that mutations in the CL domain, and CL-CH1 interface have the greatest potential for stabilisation of Fab A33. Several key salt bridge losses underpinned the conformational change in CL at low pH, whereas at high temperature, salt bridges became more dynamic, thus contributing to an overall destabilization. Lastly, the unfolding events at the two stress conditions exposed different predicted aggregation-prone regions (APR) to solvent, which would potentially lead to different aggregation mechanisms. Overall, our results identified the early stages of unfolding and stability-limiting regions of Fab A33, and the VH and CL domains as interesting future targets for engineering stability to both pH- and thermal-stresses simultaneously.

【 授权许可】

Unknown