A bridging water anchors the tethered 5-(3-aminopropyl)-2′- deoxyuridine amine in the DNA major groove proximate to the N+2 C·G base pair: Implications for formation of interstrand 5′-GNC-3′ cross-links by nitrogen mustards

Site-specific insertion of 5-(3-aminopropyl)-2′-deoxyuridine (Z3dU) and 7-deaza-dG into the Dickerson-Drew dodecamers 5′-d(C¹G ²C³G⁴A⁵A⁶T ⁷T⁸C⁹Z¹⁰C¹¹G ¹²)-3′·5′-d(C¹³G¹⁴C ¹⁵G¹⁶A¹⁷A¹⁸T¹⁹T ²⁰C²¹Z²²C²³G²⁴)-3′ (named DDD^Z10) and 5′-d(C¹G²C ³G⁴A⁵A⁶T⁷X ⁸C⁹Z¹⁰C¹¹G¹²)- 3′·5′-d(C¹³G¹⁴C¹⁵G ¹⁶A¹⁷A¹⁸T¹⁹X²⁰C ²¹Z²²C²³G²⁴)-3′ (named DDD^2+Z10) (X = Z3dU; Z = 7-deaza-dG) suggests a mechanism underlying the formation of interstrand N+2 DNA cross-links by nitrogen mustards, e.g., melphalan and mechlorethamine. Analysis of the DDD^2+Z10 duplex reveals that the tethered cations at base pairs A⁵·X ²⁰ and X⁸·A¹⁷ extend within the major groove in the 3′-direction, toward conserved Mg²⁺ binding sites located adjacent to N+2 base pairs C³·Z²² and Z¹⁰·C¹⁵. Bridging waters located between the tethered amines and either Z¹⁰ or Z²² O⁶ stabilize the tethered cations and allow interactions with the N + 2 base pairs without DNA bending. Incorporation of 7-deaza-dG into the DDD^2+Z10 duplex weakens but does not eliminate electrostatic interactions between tethered amines and Z¹⁰ O⁶ and Z²² O ⁶. The results suggest a mechanism by which tethered N7-dG aziridinium ions, the active species involved in formation of interstrand 5′-GNC-3′ cross-links by nitrogen mustards, modify the electrostatics of the major groove and position the aziridinium ions proximate to the major groove edge of the N+2 C·G base pair, facilitating interstrand cross-linking.