Asymmetry in the recA protein-DNA filament

Scott D. Lauder, Stephen C. Kowalczykowski

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

The apparent DNA site size obtained from an assay monitoring the ATPase activity of Escherichia coli recA protein (n = 3.5) differs from that determined from a direct DNA binding assay (n = 7) done under identical conditions. Investigation of this discrepancy indicates that at a DNA:protein ratio of 3.5:1, one-half of the recA protein population is less sensitive to ATPase activity inhibition by the nonhydrolyzable ATP analogue adenosine 5′-O-(3-thiotriphosphate) (ATPγS), suggesting that the recA protein filament is asymmetric with respect to NTP affinity. This asymmetry does not depend on the presence of ATPγS since the apparent Km for ATP derived from single-stranded DNA-dependent ATP hydrolysis activity is dependent on the DNA:protein ratio. Three models are proposed to account for the observed site size discrepancy and the NTP binding affinity asymmetry. They differ mainly in the intrinsic site size for each recA protein monomer and in the number of DNA-binding sites/recA molecule. Gel filtration of recA-single-stranded DNA complexes at different DNA:protein ratios complements the enzymological data and provides an additional method of distinguishing among the proposed models. The phenomenon of subunit nonequivalence within the recA protein presynaptic filament may provide a molecular basis for understanding how recA protein can discriminate between different DNA molecules during homologous pairing.

Original languageEnglish (US)
Pages (from-to)5450-5458
Number of pages9
JournalJournal of Biological Chemistry
Volume266
Issue number9
StatePublished - 1991
Externally publishedYes

Fingerprint

Rec A Recombinases
DNA
Adenosine Triphosphate
Single-Stranded DNA
Adenosine Triphosphatases
Assays
Proteins
Molecules
Escherichia coli Proteins
Escherichia coli
Gel Chromatography
Hydrolysis
Complement System Proteins
Monomers
Gels
Binding Sites
Monitoring

ASJC Scopus subject areas

  • Biochemistry

Cite this

Lauder, S. D., & Kowalczykowski, S. C. (1991). Asymmetry in the recA protein-DNA filament. Journal of Biological Chemistry, 266(9), 5450-5458.

Asymmetry in the recA protein-DNA filament. / Lauder, Scott D.; Kowalczykowski, Stephen C.

In: Journal of Biological Chemistry, Vol. 266, No. 9, 1991, p. 5450-5458.

Research output: Contribution to journalArticle

Lauder, SD & Kowalczykowski, SC 1991, 'Asymmetry in the recA protein-DNA filament', Journal of Biological Chemistry, vol. 266, no. 9, pp. 5450-5458.
Lauder SD, Kowalczykowski SC. Asymmetry in the recA protein-DNA filament. Journal of Biological Chemistry. 1991;266(9):5450-5458.
Lauder, Scott D. ; Kowalczykowski, Stephen C. / Asymmetry in the recA protein-DNA filament. In: Journal of Biological Chemistry. 1991 ; Vol. 266, No. 9. pp. 5450-5458.
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