Molecular Dynamics Modeling Based Investigation of the Effect of Freezing Rate on Lysozyme Stability

Purpose The stability of protein drug products frozen during fill finish operations is greatly affected by the freezing rate applied. Non-optimal freezing rates may lead to the denaturation of protein’s complex macromolecular conformation. However, limited work has been done to address the effect of...

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Bibliographic Details
Published in:Pharmaceutical research 2022-10, Vol.39 (10), p.2585-2596
Main Authors: Duran, Tibo, Minatovicz, Bruna, Bellucci, Ryan, Bai, Jun, Chaudhuri, Bodhisattwa
Format: Article
Language:English
Subjects:
Biochemistry
Biomedical and Life Sciences
Biomedical Engineering and Bioengineering
Biomedicine
Computer applications
Denaturation
Freezing
Ice
Investigations
Lysozyme
Macromolecules
Medical Law
Molecular dynamics
Original Research Article
Pharmacology/Toxicology
Pharmacy
Protein denaturation
Protein seeding
Protein structure
Proteins
Secondary structure
Simulation methods
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Summary:Purpose The stability of protein drug products frozen during fill finish operations is greatly affected by the freezing rate applied. Non-optimal freezing rates may lead to the denaturation of protein’s complex macromolecular conformation. However, limited work has been done to address the effect of different freezing rates on protein stability at nano-scale level. Methods The stability of a model protein, lysozyme, was investigated at atomic and molecular scale under varying freezing rates and moving ice-water interface. Ice seeding approach was adopted to initiate ice formation in this present simulation. Results The faster freezing rate (11–12 K/490 ns) applied resulted in overall smaller ice fraction within the simulation box with a larger freeze-concentrated liquid (FCL) region. Consequently, the faster freezing rate better maintained protein stability with less secondary structure deviations, higher hydration level and structural compactness, and less fluctuations at individual residues than observed following slow (5–6 K/490 ns) and medium (7–8 K/490 ns) freezing rates. The present study also identified the residues near and within helices 3, 6, 7, and 8 dominate the structural instability of the lysozyme at 247 K freezing temperature. Conclusions For the first time, ice formation in therapeutic protein solution was studied “non-isothermally” at different freezing rates using molecular dynamics simulations. Thus, a good understanding of freezing rates on protein instability was revealed by applying the developed computational model.
ISSN:0724-8741
1573-904X
DOI:10.1007/s11095-022-03358-z