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Crystal structure of the human O(6)-alkylguanine-DNA alkyltransferase
J E Wibley1, A E Pegg, P C Moody
1Department of Biochemistry, University of Leicester, University Road, Leicester LE1 7RH, UK.
Abstract:
The mutagenic and carcinogenic effects of simple alkylating agents are mainly due to O(6)-alkylation of guanine in DNA. This lesion results in transition mutations. In both prokaryotic and eukaryotic cells, repair is effected by direct reversal of the damage by a suicide protein, O(6)-alkylguanine-DNA alkyltransferase. The alkyltransferase removes the alkyl group to one of its own cysteine residues. However, this mechanism for preserving genomic integrity limits the effectiveness of certain alkylating anticancer agents. A high level of the alkyltransferase in many tumour cells renders them resistant to such drugs. Here we report the X-ray structure of the human alkyltransferase solved using the technique of multiple wavelength anomalous dispersion. This structure explains the markedly different specificities towards various O(6)-alkyl lesions and inhibitors when compared with the Escherichia coli protein (for which the structure has already been determined). It is also used to interpret the behaviour of certain mutant alkyltransferases to enhance biochemical understanding of the protein. Further examination of the various models proposed for DNA binding is also permitted. This structure may be useful for the design and refinement of drugs as chemoenhancers of alkylating agent chemotherapy.
Insights
Human O(6)-alkylguanine-DNA alkyltransferase repairs DNA damage from alkylating agents. Its structure reveals drug resistance mechanisms and aids in designing new cancer therapies.
Area of Science:
- Molecular Biology
- Biochemistry
- Structural Biology
Background:
- Simple alkylating agents cause mutations and cancer via O(6)-alkylation of guanine in DNA.
- DNA repair by O(6)-alkylguanine-DNA alkyltransferase (AGT) directly reverses this damage.
- High AGT levels in tumor cells confer resistance to alkylating anticancer drugs.
Purpose of the Study:
- To determine the X-ray structure of human AGT.
- To understand the structural basis for AGT's specificity towards O(6)-alkyl lesions and inhibitors.
- To provide insights for designing improved anticancer agents.
Main Methods:
- X-ray crystallography
- Multiple wavelength anomalous dispersion (MAD) phasing
Main Results:
- The X-ray structure of human AGT was solved.
- The structure explains differences in specificity compared to E. coli AGT.
- The structure aids in interpreting mutant AGT behavior and DNA binding models.
Conclusions:
- The human AGT structure elucidates mechanisms of drug resistance.
- This structural information can guide the development of novel chemotherapy strategies.
- Understanding AGT is crucial for enhancing alkylating agent efficacy in cancer treatment.