Mg-dependent Structural Analysis of Kink Turns in Messenger RNA L30e
Mentor: Dr. Susan White
The Saccharomyces cerevisiae (yeast) ribosomal protein L30e plays a key part in the regulation of splicing in the nucleus. It is closely related to the human L30e. The yeast L30e binds to its transcript RNA if there is excess L30e and interferes with spliceosome formation. (1)
In order to learn more yeast L30e, mutational analysis will be done. The gene will be expressed in E. coli through in vitro transcription. The focus of the research will be on the kink turn of L30e.
Kink turns are structural motifs in RNA; they provide the structural interactions needed for gene interaction. It has a conical and non-conical side with a consensus sequence, also called a conserved sequence motif.
Figure 1. A Genetic Kink Turn in which C helix is the conical and NC is non-conical helix (2)
Kink turn motifs will be inserted into extensively studied RNA in order to create various structures, such as a kink-turn RNA to compare to a base-paired RNA for us in binding experiments with magnesium. Another mutation, with the characteristics akin to both, will be attempted.
Previous experiments have shown that RNA does not form kink turns when magnesium is not present. One of the experiments to be performed is sedimentation velocity, which provides information about the molecular mass and shape, detecting changes in conformations and formations. Part of the summer will be trying to reduce unfavorable interactions like dimerization that will skew the data.
1. Kawaguchi, Akiko, Toyoyuki Ose, Min Yao, and Isao Tanaka. "Crystallization and Preliminary X-ray Structure Analysis of Human Ribosomal Protein L30e." Structural Communications F (2011): 1516-518.
2. Lilley, David M.J. "Introduction of Kink Turns in RNA." Kink-turns in RNA. University of Dundee, n.d. Web. 10 June 2013.