DIRECT OBSERVATION OF 2 BASE-PAIRING MODES OF A CYTOSINE-THYMINE ANALOG WITH GUANINE IN A DNA Z-FORM DUPLEX - SIGNIFICANCE FOR BASE ANALOG MUTAGENESIS

The pyrimidine nucleobase analogue 6H,8H-3,4-dihydropyrimido[4,5-c][1, 2]oxazin-7-one (P) is a mimic both of cytosine and thymine, since it c an form stable hydrogen-bonded base-pairs with either guanine or adeni ne. To investigate the geometric properties of pairing with guanine in a DNA double helix, the structure of d(CGCGPG)2 has been determined b y single crystal X-ray analysis. The oligonucleotide crystallised as a left-handed Z-DNA duplex in the orthorhombic space group P2(1)2(1)2(1 ) With cell dimensions a = 18.23 Angstrom, b = 30.63 Angstrom, c = 43. 78 Angstrom. Refinement using NUCLSQ with 51 water molecules included in the final model converged at R = 0.179 (R(w) = 0.159) for 2798 refl ections (F > 2 sigma(F)) in the range 8 Angstrom to 1.7 Angstrom. Rema rkably, the two P . G pairs in the hexamer duplex are different: Watso n-Crick and wobble types separately illustrate both cytosine-like and thymine-like behaviour. The result suggests that mutagenesis experimen ts involving P and other analogues which display pronounced base-pairi ng ambivalence can be used to examine the structural basis of substrat e discrimination by polymerases that is essential to accurate genetic replication. (C) 1995 Academic Press Limited