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Functional evaluation of candidate ice structuring proteins using cell-free expression systems

Paper ID Volume ID Publish Year Pages File Format Full-Text
23602 43456 2013 10 PDF Available
Title
Functional evaluation of candidate ice structuring proteins using cell-free expression systems
Abstract

Ice structuring proteins (ISPs) protect organisms from damage or death by freezing. They depress the non-equilibrium freezing point of water and prevent recrystallization, probably by binding to the surface of ice crystals. Many ISPs have been described and it is likely that many more exist in nature that have not yet been identified. ISPs come in many forms and thus cannot be reliably identified by their structure or consensus ice-binding motifs. Recombinant protein expression is the gold standard for proving the activity of a candidate ISP. Among existing expression systems, cell-free protein expression is the simplest and gives the fastest access to the protein of interest, but selection of the appropriate cell-free expression system is crucial for functionality. Here we describe cell-free expression methods for three ISPs that differ widely in structure and glycosylation status from three organisms: a fish (Macrozoarces americanus), an insect (Dendroides canadensis) and an alga (Chlamydomonas sp. CCMP681). We use both prokaryotic and eukaryotic expression systems for the production of ISPs. An ice recrystallization inhibition assay is used to test functionality. The techniques described here should improve the success of cell-free expression of ISPs in future applications.

► We describe cell-free expression methods for three ISPs that differ widely in structure and glycosylation status. ► We use both prokaryotic and eukaryotic expression systems for the production of ISPs. ► An ice recrystallization inhibition assay is used to test functionality. ► The techniques described here should improve the success of cell-free expression of ISPs in future applications.

Keywords
Ice structuring protein; Antifreeze protein; Ice binding protein; Cell-free protein synthesis; In vitro translation
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Functional evaluation of candidate ice structuring proteins using cell-free expression systems
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 163, Issue 3, 10 February 2013, Pages 301–310
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us