Evaluation of the temperature sensitivity of different Symbiodinium clades via site directed mutagenesis of glyceraldehyde-3‐phosphate dehydrogenase
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Coral bleaching occurs when corals expel their symbiotic algae (Symbiodinium). This happens especially at high temperatures, and can be fatal to the coral. Different clades of Symbiodinium, however, seem to provide different levels of thermotolerance to the holobiont, although the cause is unknown. My project tested the hypothesis that an enzyme, glyceraldehyde‐3-phosphate dehydrogenase (GAPDH), from a more heat-tolerant clade (D) is better able to resist denaturation at high temperatures than the ortholog from a more heat‐sensitive clade (C). Furthermore, I tested the effects of amino acid differences between orthologs of GAPDH C & D in an attempt to determine which are responsible for the observed difference in thermal stability. Using site‐directed mutagenesis, I altered 4 potentially thermally significant amino acids and tested residual activity after heat exposure. I found that single mutants N25G, R58K, V140C, and H343Y all significantly increased the thermal stability of clade C GAPDH, pushing its thermotolerance towards that of clade D GAPDH. A V140C/R58K double mutant was the most thermally stable, surpassing clade D GAPDH. A V140C/H343Y double mutant, however, did not demonstrate any variation from clade C GAPDH. My results indicate that it may not take very many mutations in Symbiodinium proteins to alter their temperature sensitivity. This presents the possibility that the holobiont might be able to withstand global warming and increasing SSTs through selection for more thermally stable proteins within Symbiodinium, leading to reduced bleaching.
Franklin and Marshall College Archives, Undergraduate Honors Thesis 2017
- F&M Theses Collection