DNA Structure Puzzle #6 - X-Ray Diffraction from True Crystals of Oligonucleotides

Clive Delmonte clivedelmonte at c-i-delmonte.freeserve.co.uk
Sat Nov 6 12:51:46 EST 1999

Puzzle # 6    X-Ray Diffraction from True Crystals of Oligonucleotides

In Puzzles 3, 4 & 5 are listed patterns of systematic absence reported in
some hexagonal and orthorhombic lattices in nucleic acid fibres which invite
us, at the very least, and perhaps even require us (Puzzle 4), to explore
the implications of choosing the smaller unit cell which the systematic
absences allow. An extensive analysis of some 20 papers on x-ray diffraction
from nucleic acid fibres which all show these patterns of systematic absence
has been set out (Puzzle #1, Ref. 2, Chapter 7).

Exactly the same patterns of systematic absence, sometimes to be taken with
systematically weak presences, have been reported in many studies of x-ray
diffraction from true crystals of oligonucleotides.

For example, the fragment d(CCGGCGCCGG) has been reported to crystallise in
the rhombohedral space group R3 (Ref 14, Brookhaven file 1CGC). Inspection
of the structure factor file shows that systematic absences follow the
pattern (-h+k)=/=3n for l=3n, and (-h+k)=3n when l=/=3n. This pattern allows
the selection of a unit cell whose side has been reduced from 5.407 nm,
originally selected, to 5.407 nm / root 3.

Exactly the same patterns of systematic absence arise from diffraction from
crystals of d(ACCGGCGCCACA) (Ref 15) also found in space group R3.

The fragment d(CGCG) crystallises in space group P6(1)22 and was assigned
a=b=3.08 nm (Ref 16). However, the pattern of systematic absences plus
systematically weak presences shows a reduced value of a=b=1.78 nm to
conform with the structure factor file (Ref 16, page 538).

Reduced unit cells can be chosen from other space groups. For example, in a
study of GpC crystals in space group P2(1), with a=2.1224 nm and b=3.4207
nm, the pattern of systematic absences plus systematically weak presences
allows the halving of side b (Ref 17).

Of course there are studies of diffraction from nucleic acid fibres and
oligonucleotide crystals where these patterns of systematic absence and
systematically weak presences are not very evident. However, this can be
explained by the growing complexity of the fragments inducing bends in the
sequence, by greater departures from symmetry, or perhaps by more powerful
x-ray sources generating many more reflections which may obscure the
patterns, etc.

Complexity in the Sciences is usually developed from earlier, simpler models
and studies, and this is true also in oligonucleotide crystallography. This
should have been pursued using the smallest unit cells consonant with the
observed patterns within the structure factors, and not by having to fit a
pre-conceived model (the double helix) into the unit cell irrespective of
the alternatives which the actual experimental structure factors indicated.

At present, oligonucleotide crystallographers adopt very widely a range of
practices which can compromise the outcomes of their work. For example,
research groups variously over time have adopted starting coordinates for
their structures which are explicitly taken from earlier studies, themselves
derived from fibre studies which are under scrutiny in these Puzzles. Or
they adopt unit cell dimensions consistent with previous work which also
lead back to fibre studies under scrutiny in these Puzzles. Or they adopt
structure refinement algorithms (Ref 18) which, taken with their choice of
(large) unit cell dimensions, are programmed in such a way that they are
guaranteed to lead to double helical outcomes.

The choice of unit cell dimensions determines the scales on the axes of the
resulting Pattersons, Patterson difference maps, Harker sections and
electron density maps. It will not matter whether heavy atoms, MAD,
synchrotron sources, or whatever, were used in later work if the choice of
unit cell dimensions, derived from earlier studies where alternative
possibilities had been overlooked, had thereby been rendered insecure in
later studies.

This is how the entire Brookhaven nucleic acid data base has become

We should perhaps remind ourselves that we are unable to look to NMR for
support for the double helical structure of duplex DNA because the NMR
calibrations upon which distance measurements are based assume that the
double helix is the struicture of duplex DNA. This is why "spin diffusion"
is invoked so often in NMR work on oligonucleotides in order to explain
unexpectedly high signals. The structure recorded by Lee et al. (Puzzle 1)
is more compact than the double helix and certain protons really are much
closer together than the double helical model for duplex DNA predicts.

In summary, Puzzle #6 is this: If structure factor files are deconvoluted
with an unbiassed program such as SHELX using the smallest unit cells
consistent with the reported patterns of systematic absence and
systematically weak presences, will valid, alternative duplex DNA structures
consonant with the reduced cross-section of the diagram in Puzzle 1 then be
found ?


14 Double helix conformation, groove dimensions and ligand binding potential
of a G/C stretch in B-DNA; Udo Heinemann et al.; The EMBO J., Vol 11 (1992)
1931 - 1939

15 Unusual helical packing in crystals of DNA bearing a mutation hot spot;
Y. Timsit et al.; NATURE Vol 341 (1989) 459 - 462

16 A Salt-induced Conformational Change in Crystals of the Synthetic
Tetramer d(CGCG); H. R. Drew et al.; J Mol Biol Vol 125 (1978) 535 - 543

17 The Crystal and Molecular Structure of a Calcium Salt of
Guanyl-3,5-Cytidine (GpC); B.Hingerty et al.; Acta Cryst Vol B32 (1976) 2998

18 Definitions and Nomenclature of Nucleic Acid Structure Parameters; R.
Dickerson; J Mol Biol Vol 205 (1989) 787 - 791

Clive Delmonte

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