Proteomic approach for evaluating cryoprotectant formulations for enhanced post-cryopreservation recovery of yeast

Fungi have numerous potential biotechnological applications across the life sciences. To ensure these microorganisms are available for future research, it is essential that they are properly preserved to safeguard against genetic changes, cellular instability, and loss of viability. While the use of cryoprotective agents (CPA) is critical for increasing survival of the material during preservation, the wide adoption of glycerol and DMSO as CPAs may not always be ideal as fungal diversity and functionality are ever growing. Therefore, in the following work, we focused on developing robust cryopreservation formulations that can efficiently preserve fungal strains while also maximizing recovery. Here, 10 different cryopreservation formulations consisting of individual or a combination of CPAs were evaluated for their effect on the Saccharomyces cerevisiae (ATCC 7754) proteome. Spot assays were performed to study the recovery response of each formulation. Functional proteomic and KEGG pathway analyses were used to investigate the molecular mechanism of cold-stress response in S. cerevisiae. A total of 2,299 proteins were identified; depending on the formulation used, a range of 116-1,241 proteins were found to be significantly upregulated and downregulated, indicating the influence of individual formulations. To the best of our knowledge, this is the first study that uses a proteomic-based approach to investigate how different cryopreservation formulations affect key mechanisms within a model organism.