Determination of the structural role of the internal guanine-cytosine base pair in recognition of a seven-base-pair sequence cross-linked by Bizelesin

A. S. Thompson, J. Y. Fan, Daekyu Sun, M. Hansen, Laurence Hurley

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Bizelesin (formerly U77,779, The Upjohn Co.) is a bifunctional DNA cross- linking antitumor antibiotic consisting of two open-ring homologs of the (+)- CC-1065 cyclopropa[c]pyrrolo[3,2-e]indol-4(5H)-one (CPI) subunits connected by a rigid linking moiety. Previous studies have shown that Bizelesin most often forms an interstrand cross-link through the N3 of two adenines 6 base pairs (bp) apart (inclusive of the modified adenines). However, gel electrophoresis studies have also indicated that Bizelesin forms 7-bp cross- links in specific sequences. In most of these sequences the cross-linked adenines represent the only possible cross-link site (i.e., no 6-bp site is available); however, in several sequences, a 7-bp sequence is selected in overwhelming preference to a possible 6-bp sequence. In this study, we demonstrate the unique requirement for a G·C base pair within this sequence and the critical presence of the exocyclic 2-amino group of guanine. In a subsequent two-dimensional 1H-NMR study that concentrates on the 7-bp cross- link formed with the sequence 5'-TTAGTTA-3', the role of the central G·C base pairs in the formation of a 7-bp cross-link is probed. 1H-NMR analysis coupled with restrained molecular dynamics (rMD) provides evidence for distortion around the covalently modified adenines. Because of this distortion, the modified bases are twisted toward the center of the duplex adduct, effectively reducing the cross-linked distance. The rMD study also indicates that a hydrogen bond is formed between the exocyclic amine of the central guanine and the carbonyl of the ureylene linker. On the basis of the observation of the distortion in the duplex and the hydrogen bonding between the drag and DNA, it is possible to speculate on the role of the central G·C bases in this sequence preference and propose a mechanism by which Bizelesin forms a 7-bp rather than a 6-bp cross-link with this sequence.

Original languageEnglish (US)
Pages (from-to)11005-11016
Number of pages12
JournalBiochemistry
Volume34
Issue number35
DOIs
StatePublished - 1995
Externally publishedYes

Fingerprint

Cytosine
Guanine
Adenine
Base Pairing
CC 1065
Molecular dynamics
Hydrogen bonds
Nuclear magnetic resonance
DNA
Electrophoresis
Crosslinking
Amines
Drag
Molecular Dynamics Simulation
Gels
Anti-Bacterial Agents
bizelesin
Hydrogen Bonding
Hydrogen
Observation

ASJC Scopus subject areas

  • Biochemistry

Cite this

Determination of the structural role of the internal guanine-cytosine base pair in recognition of a seven-base-pair sequence cross-linked by Bizelesin. / Thompson, A. S.; Fan, J. Y.; Sun, Daekyu; Hansen, M.; Hurley, Laurence.

In: Biochemistry, Vol. 34, No. 35, 1995, p. 11005-11016.

Research output: Contribution to journalArticle

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abstract = "Bizelesin (formerly U77,779, The Upjohn Co.) is a bifunctional DNA cross- linking antitumor antibiotic consisting of two open-ring homologs of the (+)- CC-1065 cyclopropa[c]pyrrolo[3,2-e]indol-4(5H)-one (CPI) subunits connected by a rigid linking moiety. Previous studies have shown that Bizelesin most often forms an interstrand cross-link through the N3 of two adenines 6 base pairs (bp) apart (inclusive of the modified adenines). However, gel electrophoresis studies have also indicated that Bizelesin forms 7-bp cross- links in specific sequences. In most of these sequences the cross-linked adenines represent the only possible cross-link site (i.e., no 6-bp site is available); however, in several sequences, a 7-bp sequence is selected in overwhelming preference to a possible 6-bp sequence. In this study, we demonstrate the unique requirement for a G·C base pair within this sequence and the critical presence of the exocyclic 2-amino group of guanine. In a subsequent two-dimensional 1H-NMR study that concentrates on the 7-bp cross- link formed with the sequence 5'-TTAGTTA-3', the role of the central G·C base pairs in the formation of a 7-bp cross-link is probed. 1H-NMR analysis coupled with restrained molecular dynamics (rMD) provides evidence for distortion around the covalently modified adenines. Because of this distortion, the modified bases are twisted toward the center of the duplex adduct, effectively reducing the cross-linked distance. The rMD study also indicates that a hydrogen bond is formed between the exocyclic amine of the central guanine and the carbonyl of the ureylene linker. On the basis of the observation of the distortion in the duplex and the hydrogen bonding between the drag and DNA, it is possible to speculate on the role of the central G·C bases in this sequence preference and propose a mechanism by which Bizelesin forms a 7-bp rather than a 6-bp cross-link with this sequence.",
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