Ribosome structure is remarkably conserved throughout all forms of life. However, it appears that biogenesis of these macromolecular machines is remarkably distinct in E. coli and S. cerevisiae, the two most common model systems. Despite the overall differences that exist in the two biogenic pathways there appears to at least a single point of commonality, which may provide a means to link these processes. All organisms likely contain a methyltransferase that almost always modifies two adjacent adenosines into N6,N6-dimethyladenosine within the terminal loop of all small ribosomal subunit RNA; in rare cases only a single adenosine is converted. This unique conservation of an RNA modification enzyme extends to the level of mitochondria and chloroplast ribosome biogenesis, as well. In E. coli this enzyme is termed KsgA, and Dim1 in S. cerevisiae. Investigation of this family of enzymes will reveal insights into the fundamental conservation of ribosome biogenesis. The long-term objective of this project is to understand the contribution this family of enzymes makes in the process of ribosome maturation at the molecular level.
The specific aims of this project are to identify the minimal substrate for KsgA, detail the binding interaction of KsgA to 30S ribosomal subunits, and to probe the structure, function, mechanism, and regulation of the dimethylase activity of KsgA. A host of biochemical assays with KsgA, such as activity assays on reconstituted, incomplete 30S and chemical probing of the complex, will address issues of substrate recognition. Mutagenesis and x-ray crystallographic analysis of KsgA and Dim1 will be used to probe the structure and function of both proteins. Ultimately, these studies will reveal molecular details of methyltransferase reactions as well as likely provide the first evidence for a universal step in ribosome biogenesis.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM066900-03
Application #
7169911
Study Section
Physiological Chemistry Study Section (PC)
Program Officer
Rhoades, Marcus M
Project Start
2005-02-01
Project End
2009-01-31
Budget Start
2007-02-01
Budget End
2008-01-31
Support Year
3
Fiscal Year
2007
Total Cost
$250,055
Indirect Cost
Name
Virginia Commonwealth University
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
105300446
City
Richmond
State
VA
Country
United States
Zip Code
23298
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