A macromolecular ribonucleoprotein complex called the ribosome is responsible for translation of the genetic code into proteins. The overall structure of the ribosome and its catalytic mechanisms are highly conserved across the three kingdoms of life. During all stages of protein synthesis ribosomes interact with a family of conserved protein factors that hydrolyze a bound molecule of GTP through an intrinsic GTPase mechanism and release GDP and inorganic phosphate. In isolation, the GTPase factors have very low GTPase activity. However, upon binding to the ribosome activity of their GTPase centers is highly stimulated. The hydrolysis of GTP is coupled to conformational rearrangements that promote various stages of protein synthesis. The protein factors interact with an overlapping binding site on the large ribosomal subunit known as the GTPase Associated Center (GAC). The GAC is composed of two separate regions of the 23S rRNA known as the L11 stalk and the sarcin-ricin loop (SRL), which are proposed to individually contribute to the GTPase activation of the factors and the coupling of hydrolysis to the reactions of protein synthesis. In this work the role of the SRL and L11 stalk during protein synthesis have been explored. We find that the SRL is essential for the GTPase activity of elongation factor (EF) -G and the folding of 23S rRNA. Portions of the L11 stalk are dispensable for the GTPase activity of EF-G and EF-Tu; however, mutant ribosomes translate with reduced accuracy. A correlation between the position of the L11 stalk and the binding of translation factors is explored, and we suggest that the L11 stalk plays a direct role in controlling access of the factor's GTPase G domain to the SRL. We uncover a role for rRNA tertiary interactions in maintaining the architecture of the L11 stalk as well as a tertiary pathway between the SRL and L11 stalk. Additionally, we show that removal of the entire stalk inactivates ribosomes.
|Advisor:||Noller, Harry F.|
|Commitee:||Jurica, Melissa, Rexach, Michael|
|School:||University of California, Santa Cruz|
|Department:||Molecular Cell and Developmental Biology|
|School Location:||United States -- California|
|Source:||DAI-B 73/07(E), Dissertation Abstracts International|
|Keywords:||Elongation factors, GTPase, L11 stalk, Ribosomes, Translation|
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