title: Anomalous cross-linking by mechlorethamine of DNA duplexes containing C-C mismatch pairs.
abstract: Nitrogen mustards such as mechlorethamine have previously been shown to covalently cross-link DNA through the N7 position of the two guanine bases of a d[GXC].d[GYC] duplex sequence, a so-called 1,3 G-G-cross-link, when X-Y = C-G or T-A. Here, we report the formation of a new mechlorethamine cross-link with the d[GXC].d[GYC] fragment when X-Y is a C-C mismatch pair. Mechlorethamine cross-links this fragment preferentially between the two mismatched cytosine bases, rather than between the guanine bases. The cross-link also forms when one or both of the guanine bases of the d[GCC].d[GCC] fragment are replaced by N7-deazaguanine, and, more generally, forms with any C-C mismatch, regardless of the flanking base pairs. Piperidine cleavage of the cross-link species containing the d[GCC].d[GCC] sequence gives DNA fragments consistent with alkylation at the mismatched cytosine bases. We also provide evidence that the cross-link reaction occurs between the N3 atoms of the two cytosine bases by showing that the formation of the C-C cross-link is pH dependent for both mechlorethamine and chlorambucil. Dimethyl sulfate (DMS) probing of the cross-linked d[GCC].d[GCC] fragment showed that the major groove of the guanine adjacent to the C-C mismatch is still accessible to DMS. In contrast, the known minor groove binder Hoechst 33258 inhibits the cross-link formation with a C-C mismatch pair flanked by A-T base pairs. These results suggest that the C-C mismatch is cross-linked by mechlorethamine in the minor groove. Since C-C pairs may be involved in unusual secondary structures formed by the trinucleotide repeat sequence d[CCG]n, and associated with triplet repeat expansion diseases, mechlorethamine may serve as a useful probe for these structures.
authors: Department of Biochemistry and Molecular Biology and Pharmaceutical Sciences, University of Southern California, 1985 Zonal Avenue, Los Angeles, California 90033, USA.
authors: R M
authors: RM
authors: Romero
authors: M
authors: M
authors: Mitas
authors: I S
authors: IS
authors: Haworth
mesh_terms: D000477:Alkylating Agents / Q000494:pharmacology*; D000478:Alkylation; D020137:Base Pair Mismatch; D006690:Bisbenzimidazole / Q000494:pharmacology; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D004247:DNA / Q000737:chemistry* / Q000187:drug effects; D006147:Guanine / Q000031:analogs & derivatives / Q000737:chemistry / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008466:Mechlorethamine / Q000494:pharmacology*; D009690:Nucleic Acid Conformation / Q000187:drug effects; D009692:Nucleic Acid Heteroduplexes; D010880:Piperidines / Q000494:pharmacology; D013463:Sulfuric Acid Esters / Q000737:chemistry
chemical_list: D000477:Alkylating Agents; D003432:Cross-Linking Reagents; D009692:Nucleic Acid Heteroduplexes; D010880:Piperidines; D013463:Sulfuric Acid Esters; D008466:Mechlorethamine; D006147:Guanine; C032727:piperidine; D004247:DNA; C066856:7-deazaguanine; C007482:dimethyl sulfate; D006690:Bisbenzimidazole