Identification of ice-binding peptide sequences from genetically-encoded phage libraries

Abstract

Ice-binding peptides are small molecules capable of exerting control over ice nucleation. They are of interest in in areas ranging from medically important problems such as cryosurgery on tumours and preservation of transplant organs, to more concrete everyday applications such as snowmaking or de-icing of roads. In this project, we aim to identify glycopeptides capable of inducing ice nucleation. To our knowledge, no ice-nucleating peptides have yet been identified.

Taking inspiration from an approach used in the literature for the purification of antifreeze proteins from a mixed solution, we developed a phage-display technique allowing for the selection of ice-binding peptides from a naïve library. In this method, a test tube chilled to -25°C is placed into a phage-containing solution. Ice builds around the test tube, integrating phage that display ice-binding peptides as it grows. Through multiple freezing rounds, we are able to narrow the selected population from 10^12 pfu to 10^2 pfu in five rounds. We present preliminary selection results from the SXCX3C library. Through these results, we wish to demonstrate a selection method applicable in a challenging system where the target (ice) must be able to be distinguished from the surrounding liquid. In addition, we have constructed a freezing platform capable of validating ice-nucleating properties, and model the process with gold slides coated with alkanethiol SAMs (self-assembled monolayers). Our next steps will be to synthesize and evaluate the ice-nucleating properties of peptide hits to identify the peptides that exhibit a statistically significant influence on ice nucleation temperature.

* Indicates faculty mentor.