Showcase of Undergraduate Research and Creative Endeavors (SOURCE)

Temperature Optimization, Expression, and Purification of CAIN55

CAIN55 is a gene identified from a bacteriophage that has been shown to be cytotoxic to mycobacterial species. The mycobacteriophage Cain, which acquires a total of 102 genes, is a member of the Siphoviridae family. Homology modeling of the translated amino acid sequence suggests that CAIN55 may be a cellobiohydrolase. However, homology models showing CAIN55 to be a cellobiohydrolase have generally low confidence scores, leading us to believe that this result may be incorrect. Online programs such as Raptorx and Phyre2 were used to generate homology models for this project. CAIN55 has also been proven to bind to NusA, which is a transcription regulator. Uncovering the structure of CAIN55 may be able to tell us more about CAIN55’s interaction with NusA, and thus, how CAIN55 helps regulate bacterial transcription. In this work, we have determined the optimum temperature for the expression of recombinant CAIN55 in Escherichia coli as well as attempted to purify the resulting recombinant protein from E. coli cultures using methods such as MCAC (Metal Chelate Affinity Chromatography). The purified protein will be used to determine the structure of CAIN55 by x-ray crystallography. Determining the structure of CAIN55 will be crucial for determining the function of the CAIN55.