The KsgA methyltransferase: Characterization of a universally conserved protein involved in robosome biogenesis

Publication Year:
2007
Usage 28
Downloads 17
Abstract Views 11
Repository URL:
https://scholarscompass.vcu.edu/etd/962
Author(s):
O'Farrell, Heather Colleen
Tags:
RNA modification; crystal structure; Biochemistry, Biophysics, and Structural Biology; Life Sciences
thesis / dissertation description
The KsgA enzymes comprise an ancient family of methyltransferases that are intimately involved in ribosome biogenesis. Ribosome biogenesis is a complicated process, involving numerous cleavage, base modification and assembly steps. All ribosomes share the same general architecture, with small and large subunits made up of roughly similar rRNA species and a variety of ribosomal proteins. However, the fundamental assembly process differs significantly between eukaryotes and eubacteria, not only in distribution and mechanism of modifications but also in organization of assembly steps. Despite these differences, members of the KsgA/Dim1 methyltransferase family and their resultant modification of small-subunit rRNA are found throughout evolution, and therefore were present in the last common ancestor. The first member of the family to be described, KsgA from Escherichia coli, was initially shown to be the determining factor for resistance/sensitivity to the antibiotic kasugamycin and was subsequently found to dimethylate two adenosines in 16S rRNA during maturation of the 30S subunit. Since then, numerous other members of the family have been characterized in eubacteria, eukaryotes, archaea and in eukaryotic organelles. The eukaryotic ortholog, Dim1, is essential for proper processing of the pre-rRNA, in addition to and separate from its methyltransferase function. The KsgA/Dim1 family bears sequence and structural similarity to a larger group of S-adenosyl-L-methionine dependent methyltransferases, which includes both DNA and RNA methyltransferases. In this document we report that KsgA orthologs from archaea and eukaryotes are able to complement for KsgA function in bacteria, both in vivo and in vitro. This indicates that all of these enzymes can recognize a common ribosomal substrate, and that the recognition elements must be largely unchanged since the evolutionary split between the three domains of life. We have characterized KsgA structurally, and discuss aspects of KsgA's activity in light of the structural data. We also propose a model for KsgA binding to the 30S subunit, based on solution probing data. This model sheds light on KsgA's unusual regulation and on the dual function of the Dim1 enzymes.