Department of Anesthesiology

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Mutations in the N-terminal region of RecA that disrupt the stability of free protein oligomers but not RecA-DNA complexes.

TitleMutations in the N-terminal region of RecA that disrupt the stability of free protein oligomers but not RecA-DNA complexes.
Publication TypeJournal Article
Year of Publication2000
AuthorsEldin S, Forget AL, Lindenmuth DM, Logan KM, Knight KL
JournalJ Mol Biol
Volume299
Issue1
Pagination91-101
Date Published2000 May 26
ISSN0022-2836
KeywordsAdenosine Triphosphatases, Amino Acid Substitution, Biopolymers, Catalysis, Chromatography, Gel, DNA Repair, DNA, Bacterial, DNA, Single-Stranded, DNA-Binding Proteins, Escherichia coli, Microscopy, Electron, Models, Molecular, Mutation, Protein Binding, Protein Structure, Quaternary, Radiation Tolerance, Rec A Recombinases, Static Electricity, Ultraviolet Rays
Abstract

We have introduced targeted mutations in two areas that make up part of the RecA subunit interface. In the RecA crystal structure, cross-subunit interactions are observed between the Lys6 and Asp139 side-chains, and between the Arg28 and Asn113 side-chains. Unexpectedly, we find that mutations at Lys6 and Arg28 impose sever defects on the oligomeric stability of free RecA protein, whereas mutations at Asn113 or Asp139 do not. However, Lys6 and Arg28 mutant proteins showed an apparent normal formation of RecA-DNA complexes. These results suggest that cross-subunit contacts in this region of the protein are different for free RecA protein filaments versus RecA-DNA nucleoprotein filaments. Mutant proteins with substitutions at either Lys6 or Arg28 show partial inhibition of DNA strand exchange activity, yet the mechanistic reasons for this inhibition appear to be distinct. Although Lys6 and Arg28 appear to be more important to the stability of free RecA protein, as opposed to the stability of the catalytically active nucleoprotein filament, our results support the idea that the cross-subunit interactions made by each residue play an important role in optimizing the catalytic organization of the active RecA oligomer.

DOI10.1006/jmbi.2000.3721
Alternate JournalJ. Mol. Biol.
PubMed ID10860724
Grant ListGM44772 / GM / NIGMS NIH HHS / United States