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Molecular recognition between oligopeptides and nucleic acids. Specificity of binding of a monocationic bis‐furan lexitropsin to DNA deduced from footprinting and 1 H NMR studies
Author(s) -
Lee Moses,
Krowicki Krzysztof,
Shea Regan G.,
Lown J. William,
Pon Richard T.
Publication year - 1989
Publication title -
journal of molecular recognition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.401
H-Index - 79
eISSN - 1099-1352
pISSN - 0952-3499
DOI - 10.1002/jmr.300020206
Subject(s) - footprinting , stereochemistry , chemistry , two dimensional nuclear magnetic resonance spectroscopy , base pair , recognition sequence , dna , ecori , binding site , nuclear magnetic resonance spectroscopy , crystallography , restriction enzyme , biochemistry , base sequence
MPE‐Fe(EDTA) footprinting of a novel monocationic bis‐furan lexitropsin 6 on a HindIII/EcoRI restriction fragment of pBR322 DNA revealed a series of four‐base binding sites (all 5′ → 3′) of (primary) TGTA, TGAA, AAAT, ACAA, TTAT, and (secondary) CTAA, TCGT, TGTA, GTCA, and GGTT. Thus 6 can accept a GC pair at positions 1, 2 or 3 of the binding site with a strict 3′ (4 position) AT requirement. Marked enhancement of cleavage, particularly at GC rich sequences, is observed at regions flanking or even up to 18 base pairs remote from a given binding site. The non‐exchangeable and imino 1 H NMR resonances of the 1:1 complex and d‐[CATGGCCATG] 2 were assigned using a combination of NOE differences, NOESY and COSY techniques. 1 H NMR studies (ligand induced chemical shifts and NOE differences) of Lexitropsin 6 with d‐[CATGGCCATG] 2 show unambiguously the location and orientation of the N to C termini of 6 on the sequence 5′‐G 5 C 6 C 7 A 8 ‐3′, with the C terminus oriented to A 8 . This orientation of 6 in the minor groove of 5′‐GCCA is confirmed by an NOE observed between H1 2a of 6 and AH 8 (8). This preference for binding of 6 to the sequence 5′‐GCCA when challenged with d‐[CATGGCCATG] 2 is in accord with the conclusions of the footprinting experiments wherein GC base pairs can be accepted in the first three positions and with a strict 3′ terminus AT reading requirement. Collectively the data support the inference of a GC recognizing capacity for a 2,5‐substituted furan moiety within a lexitropsin. The 1 H NMR data indicate that the decadeoxyribonucleotide duplex exists in the B conformation in both the 1:1 complex and the free from. The apparent binding constant of 6 to calf thymus DNA is 1.68 × 10 5 M −1 whereas netropsin under similar conditions gives a value of 1.85 × 10 7 M −1 . This suggests that if advantage is to be taken of the GC recognizing property of a 2,5‐substituted furan in longer lexitropsins it should be flanked by more strongly bound moieties.