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Conformational preference and ligand binding properties of DNA junctions are determined by sequence at the branch
Author(s) -
Guo Qiu,
Lu Min,
Kallenbach N. R.
Publication year - 1991
Publication title -
biopolymers
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/bip.360310402
Subject(s) - holliday junction , chemistry , stacking , footprinting , dna , crystallography , oligonucleotide , endonuclease , cleavage (geology) , stereochemistry , nucleotide , affinities , bond cleavage , dna footprinting , biophysics , dna binding protein , base sequence , biochemistry , dna repair , geotechnical engineering , organic chemistry , fracture (geology) , biology , transcription factor , engineering , gene , catalysis
Four‐arm DNA branched junctions are stable analogues of Holliday recombinational intermediates. A number of four‐arm DNA junctions synthesized from oligonucleotides have now been studied. Gel mobility or chemical footprinting experiments on several immobile four‐arm junctions indicate that in the presence of Mg 2+ , they assume a preferred conformation consisting of two helical domains, each formed by stacking a particular pair of arms on each other. We show here that a junction we designate as J1 c that has the same chemical composition as one we have previously studied in detail, J1, but is formed from the four strands complementary to those of the latter, exhibits the reverse stacking preference. The pattern of self‐protection of the strands of J1 c exposed to Fe(II) · EDTA‐induced scission reveals that twofold symmetry is preserved, but the opposite pair of strands preferentially cross over. Moreover, the Fe(II) · EDTA scission profiles of J1 c indicate that this junction exhibits a weaker bias as to which strands cross over than is observed in J1. The preference for the dominant species in J1 is 1.3 times greater than in J1 c at 4°C and in the presence of 10 m M Mg 2+ , based on chemical reactivity data. This is confirmed by a cleavage experiment using the resolvase enzyme, endonuclease I, from bacteriophage T7. This difference could reflect either sequence‐dependent differences in the equilibrium among isomers, or in the structure of these junctions. Chemical footprinting experiments using the probes MPE · Fe(II) and (OP) 2 Cu(I) show that the high‐affinity ligand binding site in immobile junctions is determined by junction geometry.