University of Louisville
Sensitive to salt? Uncovering the features of a repair protein to aid dry-preservation of biological materials
Grade Level at Time of Presentation
Sophomore
Major
Biology - Molecular, Cellular, Developmental
Minor
Chemistry and Physics
KY House District #
100
KY Senate District #
31
Faculty Advisor/ Mentor
David Grimm; Michael A. Menze, PhD
Department
Department of Biology
Abstract
Successful long-term preservation of products containing biological materials like DNA, RNA, proteins, or entire cells requires storage temperatures well below 32 °F (0 °C) and often even as low as -320 °F (-196 °C). However, maintaining ultra-low temperatures is a significant financial burden, compounded by the logistical challenges of transporting frozen samples. An alternative strategy that mitigates these disadvantages is dry-state preservation, a technique that removes water from the sample before storage. This method allows for storage at room temperature, safer handling of specimens, and may also extend the shelf-life of the materials. Maintaining viable cells in a dehydrated state has several applications including life-saving blood transfusions that can be kept longer than 42 days without refrigeration. Dry storage has been successfully implemented for relatively simple biological compounds but preserving complex systems like cells has been more challenging. We turned to nature to identify biomolecules that allow cells to survive drying and found an exceptional repair protein, p26, produced by encysted embryos of a brine shrimp that can survive dehydration (desiccation) for decades. While some work has been done to understand how this protein functions, researchers have struggled to isolate adequate amounts of the pure protein to perform robust experiments. The present work details a renewed effort on that front. We enhanced production of the protein in bacteria and discovered that it is exceptionally salt-sensitive, but yields can be improved by adding the sugar trehalose and optimizing the types of salt present during purification. Analyzing the amino acid sequence of p26 with bioinformatic programs revealed regions that are likely to undergo liquid-liquid phase separation, a property shared by other proteins known to allow cells to survive desiccation. Establishing an optimal purification method will allow us to further explore the mechanism(s) by which p26 protects desiccated cells.
Sensitive to salt? Uncovering the features of a repair protein to aid dry-preservation of biological materials
Successful long-term preservation of products containing biological materials like DNA, RNA, proteins, or entire cells requires storage temperatures well below 32 °F (0 °C) and often even as low as -320 °F (-196 °C). However, maintaining ultra-low temperatures is a significant financial burden, compounded by the logistical challenges of transporting frozen samples. An alternative strategy that mitigates these disadvantages is dry-state preservation, a technique that removes water from the sample before storage. This method allows for storage at room temperature, safer handling of specimens, and may also extend the shelf-life of the materials. Maintaining viable cells in a dehydrated state has several applications including life-saving blood transfusions that can be kept longer than 42 days without refrigeration. Dry storage has been successfully implemented for relatively simple biological compounds but preserving complex systems like cells has been more challenging. We turned to nature to identify biomolecules that allow cells to survive drying and found an exceptional repair protein, p26, produced by encysted embryos of a brine shrimp that can survive dehydration (desiccation) for decades. While some work has been done to understand how this protein functions, researchers have struggled to isolate adequate amounts of the pure protein to perform robust experiments. The present work details a renewed effort on that front. We enhanced production of the protein in bacteria and discovered that it is exceptionally salt-sensitive, but yields can be improved by adding the sugar trehalose and optimizing the types of salt present during purification. Analyzing the amino acid sequence of p26 with bioinformatic programs revealed regions that are likely to undergo liquid-liquid phase separation, a property shared by other proteins known to allow cells to survive desiccation. Establishing an optimal purification method will allow us to further explore the mechanism(s) by which p26 protects desiccated cells.