Understanding how extremophiles have evolved stable and functional proteins is difficult, as most adaptations are not obvious and/or made by the combination of different features. Halophilic proteins are no different and are still misunderstood as to how they maintain their stability. To try and understand how these proteins maintain their structure under such extreme conditions, we are studying the highly conserved protein, cysteinyl t-RNA synthetase (CysRS), which catalyzes the production of cysteinyl-tRNACys. In order to probe its structure, we are using the protein binding dye 1-anilinonaphthalene-8-sulfonate (ANS), which increases its fluorescence in the presence of a hydrophobic environment. Traditionally, this is used to indicate the degree of unfolding as ANS will bind to the exposed hydrophobic core. From previous studies, it was shown that group I metals can increase halophilic protein structural stability. To further support this hypothesis, both the H. salinarum (Hs) and E. coli CysRS were used in an ANS binding assay to investigate the structural changes associated with various salt concentrations and temperatures. From our assays, as salt concentrations increased, ANS fluorescence increased more for the Hs CysRS than for the E. coli CysRS. This suggests that the halophilic CysRS might be exposing its hydrophobic core as salt increases. By doings so, the protein may have found a way to keep ions at bay in order to stabilize the structure. This result could describe a new feature of halophilic proteins. Overall, this project seeks to determine how salt concentrations change the Hs CysRS structure, so we can draw conclusions about the interactions between the protein and the ions. These studies can give us more information on what makes halophilic proteins stable under such extremely ionic conditions and how their proteins have evolved to maintain stability.

Surveying the salt-stabilized structure of Halobacterium salinarum Cysteinyl-tRNA synthetase by ANS fluorescence

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.