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|Title:||Investigation of Ribosome Structure and Ribosome-Transfer Rna Interaction Using T4 Rna Ligase|
|Author(s):||Rose, Samuel Jahile, Iii|
|Department / Program:||Biochemistry|
|Degree Granting Institution:||University of Illinois at Urbana-Champaign|
|Abstract:||T4 RNA ligase can be used to alter the sequence of E. coli ribosomal RNAs in the intact polyribosome, ribosome and subunits. This enzyme will catalyze the addition of a 3',5'-bisphosphate onto the 3' terminus of rRNAs resulting the rRNA molecules one nucleotide longer with a 3' phosphate. The RNA ligase catalyzed addition of radioactively labeled cytidine 3',5'-bisphosphate has been used to probe the availability of the 3' ends of rRNAs in the ribosome under varying conditions.
T4 RNA ligase can also be used to synthesize the fifteen nucleotide long anticodon arm region of yeast tRNA('phe) complete with modified nucleotides and a four base pair helical stem. This anticodon arm was examined with respect to binding the 30S subunit and 70S ribosome of E. coli compared to whole yeast tRNA('phe). Equilibrium binding constants were derived under various binding conditions using two different assay methods. The effect of varying the mRNA length and sequence and varying the nucleotide sequence of the synthetic anticodon arm upon the binding constant was examined. It has been shown that an intact base paired stem is necessary for the binding of the anticodon arm to the 30S subunit.
Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1981.
|Date Available in IDEALS:||2014-12-14|