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Title:Biochemical and Structures Studies of tRNA Modificaton and Repair Enzymes
Author(s):Zhou, Chun
Doctoral Committee Chair(s):Huang, Raven H
Department / Program:Biochemistry
Discipline:Biochemistry
Degree Granting Institution:University of Illinois at Urbana-Champaign
Degree:Ph.D.
Genre:Dissertation
Subject(s):Chemistry, Biochemistry
Abstract:Plant siRNAs and miRNAs are methylated at their 3' ends at the 2' OH position by Hen1, an S-Adomet dependent methyltransferase. Hen1 homologs in animals are shown to be responsible for the methylation of piRNAs, some siRNAs as well as endogenous siRNAs. Interestingly, a subset of bacteria also has a Hen1 homolog. Since bacteria do not have canonical RNAi, Hen1 in bacteria (bHen1) might play other biological roles, Our goals are to find out the biological function of bHen1. We observed that immediately following bHen1, there was a second highly conserved gene (Pnkp) in the operon. Pnkp has previously been shown to have kinase, phosphatase and adenylyltransferase activity. Therefore Pnkp/Hen1 might be involved in RNA repair. I cloned and expressed both proteins with a pETDuet vector. Pnkp and bHen1 form a heterotetramer in gel filtration column. Purfied Pnkp/bHEN1 efficiently repaired full length tRNAs cleaved by bacterial toxins (colicin E5 and D). Before the broken RNAs were ligated, a methyl group was added to the 2' OH group that participated in the original RNA cut. Due to this methylation, RNAs repaired by bacterial Pnkp/Hen1 could not be cleaved again by the ribotoxins. Thus, unlike eukaryotic Hen1 involved in RNAi, bacterial Hen1 is part of a RNA repair and modification system.
Issue Date:2009
Type:Text
Language:English
Description:108 p.
Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2009.
URI:http://hdl.handle.net/2142/84867
Other Identifier(s):(MiAaPQ)AAI3395562
Date Available in IDEALS:2015-09-25
Date Deposited:2009


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