Abstract
Proteins are ubiquitous in molecular biotechnology, biotechnology and as therapeutics, but there are significant challenges in their storage and distribution, with freezing often required. This is traditionally achieved by the addition of cryoprotective agents such as glycerol (or trehalose) or covalent modification of mutated proteins with cryoprotectants. Here, ice recrystallization inhibiting polymers, inspired by antifreeze proteins, are used synergistically with poly(ethylene glycol) as an alternative to glycerol. The primary mechanism of action appears to be preventing irreversible aggregation due to ice growth. The polymer formulation is successfully used to cryopreserve a range of important proteins including insulin, Taq DNA polymerase and an IgG antibody. The polymers do not require covalent conjugation, nor modification of the protein and are already used in a wide range of biomedical applications, which will facilitate translation to a range of biologics.
Highlights
The current method used in biochemical, molecular biology, protein engineering and therapeutic labs for freezing proteins is often based on adding organic solvents to modulate ice formation and stabilise the proteins
We identified that irreversible protein aggregation due to ice crystal growth is a major cause of cryo-damage and that if we prevent this, the proteins retain activity
We hypothesized that ice recrystallization inhibition (IRI)-active compounds might prevent protein aggregation during ice-growth induced stress, which normally leads to denaturation/deactivation
Summary
The current method used in biochemical, molecular biology, protein engineering and therapeutic labs for freezing proteins is often based on adding organic solvents to modulate ice formation and stabilise the proteins. Which along with irreversible aggregation result in inactivated proteins.[2,8] Current solutions to this challenge include lyophilization or direct freezing in solution with the addition of large concentrations of osmolytes that make unfolding thermodynamically less favourable,[9] though more recently spray drying and vacuum foam drying have been introduced.[10,11] Whilst these methods are successful, there are post-thaw issues associated with the compatibility of high concentrations of osmolytes used (e.g. 10–20% glycerol) This can include sample viscosity or toxicity,[12] interference with colorimetric assays, affecting protein–. This effect could in turn be exploited in the solvent-free storage proteins
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
