Fluorescence-detected interactions of oligonucleotides in RecA complexes
Artikel i vetenskaplig tidskrift, 1995

A technique has been developed to probe directly RecA-DNA interactions by the use of the fluorescent chromophore, (+)anti-benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE), covalently attached to DNA, The 24-mer oligonucleotide 5'-d(CTACTAAACATGTACAAATCATCC) was specifically modified on the exocyclic nitrogen of the central guanine, to yield a trans-adduct. Upon interaction of the modified oligonucleotide with RecA we find an increase in BPDE fluorescence and a rather high fluorescence anisotropy, suggesting a restricted motion of the BPDE-oligonucleotide in the protein filament. In the presence of the cofactor ATP gamma S, binding of two oligonuclotides, identical or complementary in sequence, in the RecA filament is possible, The RecA-DNA complex is, however, more stable when the sequences are complementary; in addition, a shift in the BPDE emission peaks is observed, In the presence of ATP (and an ATP regeneration system), the RecA-DNA interaction between two complementary oligonucleotides is changed, and we now find protein-mediated renaturation to occur.

escherichia-coli

solution

stoichiometry

spectroscopy

protein

mechanism

genetic-recombination

linear dichroism

reca

coordination

benzo(a)pyrenediolepoxide

conformation

dna complexes

recombination

fluorescence

renaturation

Författare

Pernilla Wittung Stafshede

Institutionen för fysikalisk kemi

M. Funk

B. Jernstrom

Bengt Nordén

Institutionen för fysikalisk kemi

M. Takahashi

FEBS Letters

0014-5793 (ISSN)

Vol. 368 1 64-68

Ämneskategorier

Biokemi och molekylärbiologi

DOI

10.1016/0014-5793(95)00600-E