Exploring Secondary Structure In Bacteriophage Programmed Frameshift Elements
Dr. Sean McClory
Bacteriophages are viruses that infect bacteria and reproduce using host bacterial components. Part of the bacteriophage reproduction is assembly of the tail complex, which requires two assembly chaperone (TAC) proteins. In many phages the TAC’s are produced from a single gene through a non-canonical process called programmed translational frameshifting (PTF). The SEA-PHAGES program has produced hundreds of TAC genes that are accessible through phagesdb, a database of sequenced and annotated phage genomes. The sequences for the TAC gene were gathered from phagesdb and analyzed using ClustalOmega; a multiple sequence alignment (MSA) tool which revealed several positions where total conservation was achieved over 20 different TAC genes. mFold was used to visualize the secondary structure elements within the TAC genes. Several variations of stem-loop structures were repeated and the data from the MSA analysis was used to identify conserved areas within the secondary structures. To test if PTF is occurring within the TAC genes, a reporter system must be designed so that a specific region of the TAC gene (containing the slippery sequence and downstream secondary elements) can be excised and inserted into a dual-luciferase plasmid reporter. If PTF is occurring, the dual-luciferase protein will be translated successfully, translation of only one of the luciferase proteins shows that PTF is not occurring. Current work with the dual-luciferase reporter has shown that an insert was successful, but the sequence of interest was inserted backwards.
Okabayashi, Samuel and McClory, Sean, "Exploring Secondary Structure In Bacteriophage Programmed Frameshift Elements" (2019). HON499 projects. 33.