Effects of a carcinogenic alkyl lesion on DNA: A structural study by x-ray crystallography
Mahalingam, Sriram
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https://hdl.handle.net/2142/19467
Description
Title
Effects of a carcinogenic alkyl lesion on DNA: A structural study by x-ray crystallography
Author(s)
Mahalingam, Sriram
Issue Date
1994
Doctoral Committee Chair(s)
Wang, Andrew H.J.
Department of Study
Biophysics
Discipline
Biophysics
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Molecular
Biophysics, General
Language
eng
Abstract
"$\rm O\sp6$-ethyl-G $\rm (e\sp6G)$ is a persistent alkylation lesion of DNA, caused by the exposure of cells to alkylating agents such as N-ethyl-N-nitrosourea. Alkylating N-nitroso compounds are extremely significant as environmental carcinogens. It has been established experimentally that $\rm O\sp6$-alkyl-guanine $\rm (a\sp6G)$ can produce point mutations in vitro and in vivo because of its miscoding behavior. Persistent $\rm a\sp6G$ adducts have been proposed to be the critical lesion in a variety of alkylating agent-induced experimental tumors, such as mouse lymphomas and rat mammary tumors, because these tumors frequently display G$\to$A activating mutations in K-ras or H-ras. In this work, the three-dimensional structure of a DNA molecule incorporating the $\rm e\sp6G$ lesion has been determined by X-ray crystallography. The DNA dodecamer d(CGC ($\rm e\sp6G$) AATTCGCG) could be crystallized only when complexed to minor groove binding drugs like Hoechst 33258, Hoechst 33342, Netropsin, as well as SN6999. In addition, the native dodecamer d(CGCGAATTCGCG) was crystallized with three drugs for comparison. The structures were solved by molecular replacement method and refined by constrained least squares procedure to R-factors of $\sim$16% at $\sim$2.5 A resolution. The two independent $\rm e\sp6G$:C base pairs in the DNA duplex adopt different base pairing schemes. A bifurcated base-pair configuration of $\rm e\sp6G$:C with three-centered hydrogen bonds mimicking a Watson-Crick pairing, a wobble configuration, as well as a ""reverse wobble"" configuration with cytosine pushed towards the major groove side, were observed. A dynamic equilibrium between these configurations for the $\rm e\sp6G$:C base pair is likely and would present an ambiguous signal to the cellular replication or repair mechanisms. The results suggest that the $\rm e\sp6G$:C base pairing is weak and polymorphic when compared to a normal G:C base-pair and the DNA duplex containing this lesion is readily distorted."
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