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Propidium binding to a ribonuclease-DNA complex: X-ray and fluorescence studies
M McGrath1, D Cascio, R Williams
1Department of Biochemistry, University of California, Riverside 92521.
Molecular Pharmacology
|November 1, 1987
Summary
Propidium iodide, an antitumor drug, binds to RNase A near methionine 13 and phenylalanine 8, not the active site. This binding competitively inhibits RNase A activity, affecting nucleotide binding.
Area of Science:
- Biochemistry
- Structural Biology
- Enzymology
Background:
- RNase A is an enzyme that degrades RNA.
- Propidium iodide is an antitumor compound with potential enzyme inhibitory properties.
- Understanding drug-protein interactions is crucial for therapeutic development.
Purpose of the Study:
- To investigate the binding site and mechanism of propidium iodide with RNase A.
- To determine the effect of propidium iodide binding on RNase A's enzymatic activity.
- To characterize propidium iodide as a potential inhibitor of RNase A.
Main Methods:
- Crystallization of RNase A complexed with deoxytetraadenylate (dpA)4.
- X-ray crystallography (difference Fourier analysis at 2.9 A) to determine the binding site.
- Fluorescence titrations to assess inhibitory activity and binding kinetics.
Main Results:
- Propidium iodide binds to a hydrophobic depression on RNase A, involving methionine 13 and phenylalanine 8, away from the active site.
- Propidium binding induces minor conformational changes affecting nucleotide binding at the active site.
- Propidium iodide acts as a competitive inhibitor of RNase A with a Ki of approximately 1 mM.
Conclusions:
- Propidium iodide inhibits RNase A through competitive inhibition at a non-active site.
- The binding site and conformational changes suggest a novel mechanism for enzyme inhibition.
- This study provides insights into the interaction of propidium iodide with RNase A, relevant for antitumor drug design.