Ribonucleoprotein (RNP) complexes are integral to cellular function and control of RNP assembly is an important cellular regulatory mechanism and an attractive therapeutic target. One interesting antibiotic target is the RNase P complex that is responsible for 5'-tRNA maturation and, thus, integral to protein synthesis. RNase P is also a tractable system for investigating RNP assembly since in Bacillus subtilis it is composed of a single RNA and a relatively small protein subunit. This proposal addresses experiments designed to gain a more detailed understanding of RNP assembly by investigating the mechanism of RNase P protein folding. Thermodynamic studies using small anionic ligands indicate that P protein folding is a two-state process in which binding and folding are coupled. Binding stoichiometries clearly indicate that two specific sites are responsible for small anion binding/folding. Photoaffinity labeling, mutagenesis and NMR will be used to identify and characterize these specific binding sites and to assess their role in P RNA binding. In addition, amide exchange, 15N relaxation and heteronuclear NMR experiments will be used to characterize changes in conformational dynamics and structure as a function of ligand (small anion vs. RNA). Finally, the kinetic folding pathway for P protein will be determined to gain insight into the requirements for folding in vivo.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32GM066505-02
Application #
6646521
Study Section
Special Emphasis Panel (ZRG1-F04 (20))
Program Officer
Cassatt, James
Project Start
2002-09-01
Project End
2005-11-15
Budget Start
2003-09-01
Budget End
2004-11-15
Support Year
2
Fiscal Year
2003
Total Cost
$41,608
Indirect Cost
Name
Duke University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
044387793
City
Durham
State
NC
Country
United States
Zip Code
27705