Regioregular Alternating Polyampholytes Have Enhanced Biomimetic Ice Recrystallization Activity Compared to Random Copolymers and the Role of Side Chain versus Main Chain Hydrophobicity.

Christopher Stubbs,Julia Lipecki,Matthew I Gibson

doi:10.1021/acs.biomac.6b01691

Christopher Stubbs, Julia Lipecki + Show 1 more

Open Access

https://doi.org/10.1021/acs.biomac.6b01691

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Journal: Biomacromolecules	Publication Date: Dec 23, 2016
Citations: 57	License type: cc-by

Affiliation: University of Warwick

Abstract

Antifreeze proteins from polar fish species are potent ice recrystallization inhibitors (IRIs) effectively stopping all ice growth. Additives that have IRI activity have been shown to enhance cellular cryopreservation with potential to improve the distribution of donor cells and tissue. Polyampholytes, polymers with both anionic and cationic side chains, are a rapidly emerging class of polymer cryoprotectants, but their mode of action and the structural features essential for activity are not clear. Here regioregular polyampholytes are synthesized from maleic anhydride copolymers to enable stoichiometric installation of the charged groups, ensuring regioregularity, which is not possible using conventional random copolymerization. A modular synthetic strategy is employed to enable the backbone and side chain hydrophobicity to be varied, with side chain hydrophobicity found to have a profound effect on the IRI activity. The activity of the regioregular polymers was found to be superior to those derived from a standard random copolymerization with statistical incorporation of monomers, demonstrating that sequence composition is crucial to the activity of IRI active polyampholytes.

Highlights

Antifreeze proteins (AF(G)Ps) have evolved in polar fish species to enable them to survive in subzero environments by interacting with ice crystals.[1]
AF(G)Ps have three main properties of thermal hysteresis, dynamic ice shaping, and ice recrystallization inhibition (IRI), the inhibition of the growth of already formed ice crystals.[1−3] The property of ice recrystallization inhibitors (IRIs) is of particular interest as the growth of ice crystals has been identified as a crucial mechanism of cell death during the thawing of cryopreserved cells and tissues.[4]
To enable the first detailed study on structure−activity relationships in well-defined and regioregular polyampholytes, copolymers based on maleic anhydride were designed to give a perfectly alternating structure.[39]

Summary

Introduction

Antifreeze (glyco) proteins (AF(G)Ps) have evolved in polar fish species to enable them to survive in subzero environments by interacting with ice crystals.[1] AF(G)Ps have three main properties of thermal hysteresis (lowering freezing point, but not melting point), dynamic ice shaping (changing morphology of ice crystals), and ice recrystallization inhibition (IRI), the inhibition of the growth of already formed ice crystals.[1−3] The property of IRI is of particular interest as the growth of ice crystals has been identified as a crucial mechanism of cell death during the thawing of cryopreserved cells and tissues.[4] The addition of AF(G)Ps to cryopreservation solutions was found to give some increase in cell recovery but the effect was limited by the onset of dynamic ice shaping, which leads to ice crystals piercing cell membranes.[5]. Effective cryoprotectants are important for applications in frozen food.[12−14] Current cryopreservation strategies often involve addition of large quantities of DMSO which can have negative (toxic) effects on both the cells and the recipient.[15,16]

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