文章基本信息

标题：Comparison of the pH- and thermally-induced fluctuations of a therapeutic antibody Fab fragment by molecular dynamics simulation
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作者：Cheng Zhang ; Nuria Codina ; Jiazhi Tang 等
期刊名称：Computational and Structural Biotechnology Journal
印刷版ISSN：2001-0370
出版年度：2021
卷号：19
页码：2726-2741
DOI：10.1016/j.csbj.2021.05.005
出版社：Computational and Structural Biotechnology Journal
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 C L . 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 V L -V H and C L -C H 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 C L domain, and C L -C H 1 interface have the greatest potential for stabilisation of Fab A33. Several key salt bridge losses underpinned the conformational change in C L 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 V H and C L domains as interesting future targets for engineering stability to both pH- and thermal-stresses simultaneously.
关键词：Antibody fragment ; Crystal structure ; Protein stability ; Protein aggregation ; Protein engineering ; Molecular dynamics simulations