Direct observation of two base-pairing modes of a cytosine-thymine analogue with guanine in a DNA z-form duplex: significance for base analogue mutagenesis. [Dataset]

Abstract

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 can form stable hydrogen-bonded base-pairs with either guanine or adenine. To investigate the geometric properties of pairing with guanine in a DNA double helix, the structure of d(CGCGPG)2 has been determined by single crystal X-ray analysis. The oligonucleotide crystallised as a left-handed Z-DNA duplex in the orthorhombic space group P212121 with cell dimensions a = 18.23 Å, b = 30.63 Å, c = 43.78 Å. Refinement using NUCLSQ with 51 water molecules included in the final model converged at R = 0.179 (Rw = 0.159) for 2798 reflections (F > 2s(F)) in the range 8 Å to 1.7 Å. Remarkably, the two P·G pairs in the hexamer duplex are different: UK Watson-Crick and wobble types separately illustrate both cytosine-like and thymine-like behaviour. The result suggests that mutagenesis experiments involving P and other analogues which display pronounced base-pairing ambivalence can be used to examine the structural basis of substrate discrimination by polymerases that is essential to accurate genetic replication. The data for this output can be found at : https://doi.org/10.2210/pdb223d/pdb.

Citation

MOORE, M.H., VAN MEERVELT, L., SALISBURY, S.A., KONG THOO LIN, P. and BROWN, D.M. 1995. Direct observation of two base-pairing modes of a cytosine-thymine analogue with guanine in a DNA z-form duplex: significance for base analogue mutagenesis. [Dataset]. Hosted on Worldwide Protein Data Bank (wwPDB) [online], 223D. Available from: https://doi.org/10.2210/pdb223d/pdb

• Acceptance Date: Jun 27, 1995
• Online Publication Date: May 25, 1995
• Publication Date: Aug 1, 1995
• Deposit Date: Aug 23, 2022
• Publicly Available Date: Oct 27, 2022
• Publisher: Worldwide Protein Data Bank
• DOI: https://doi.org/10.2210/pdb223d/pdb
• Keywords: Pyrimidine analogue; P; Mutagenesis; crystallography; P·G base-pairs
• Public URL: https://rgu-repository.worktribe.com/output/1740183
• Collection Date: Dec 9, 1995
• Collection Method: The monomer, 5'-O-dimethoxytrityl-2'-deoxy-d-ribofuranosyl-(6H,8H-3,4-dihydropyrimido [4,5-c][1,2]oxazin-7-one)-3'-O-(2-cyanoethyl)-N,N-di-isopropyl phosphoramidite, was prepared as described (Kong Thoo Lin & Brown, 1994). The oligomer d(CGCGPG) was synthesised by automated phosphoramidite chemistry on an Applied Biosystems 381A Synthesiser and purified by ion exchange liquid chromatography and reverse phase high pressure liquid chromatography. Crystals were grown at 4 C from a solution of 2 mM oligonucleotide, 25 mM sodium cacodylate buffer (pH 6.5) and 15 mM MgCl2 equilibrated against 50% (v/v) 2-methyl-2,4-pentanediol by vapour diffusion. Intensities from a single crystal (0.25 mm 0.3 mm 0.4 mm) mounted in a glass capilliary and kept at a temperature of 19 C were measured using an AFC5R Rigaku diffractometer and a Rigaku RU200HB copper rotating anode source. Accurate unit cell dimensions, a = 18.23 Å, b = 30.36 Å, c = 43.78 Å, were obtained by least squares refinement of the values of 25 centred reflections in the range 15 < 2Ө < 28 , space group P212121. Two octants (hkl, hk l) were collected and merged using the data reduction part of the TEXSAN software to give 2958 unique reflections (Rsymm = 4%). The intensities were corrected for Lorentz and polarization effects, absorption and decay.