Other Information in Diffraction Patterns

The classic example is monoclinic C2 horse hemoglobin studied by Max Perutz. In this crystal form it was known from density measurements that the C2 unit cell contained two entire molecules of hemoglobin. There are four asymmetric units in a C2 unit cell, however, as the International Tables show. Thus one-half of a hemoglobin, to satisfy space group symmetry, had to be related to the other half by a twofold axis. By this means horse hemoglobin was shown to possess a perfect twofold axis of symmetry well before it was demonstrated by any other means. 

If one can measure, usually on some sort of mixed fluid gradient, the density of a protein crystal, then the number of molecules in the unit cell can be calculated from the unit cell volume (see McPherson, 1982,1999). The amount of protein (or number of subunits) in the asymmetric unit can then be obtained from dividing by the number of asymmetric units in the cell (known from the space group). 

A more convenient method that does not require direct measurement of the crystal density was introduced by Matthews (1968), though one must be cautious in its application. It has occasionally proved misleading, particularly for crystals of unusually high solvent content. Matthews pointed out (and more recent data seem to confirm it) that the ratio between the volume of the asymmetric unit (the volume of the unit cell divided by the number of 

For oligomeric proteins the mass of protein in the asymmetric unit is essentially quantized that is, it is always some integral multiple of the subunit molecular weight. Given 

FIGURE 6.22 Diffraction images frequently reveal problems with particular crystals that are sometimes blatant but occasionally subtle. These include disorder, multiple crystals, or twinned crystals. In (a), the pattern initially appears very ordered and proper, but close inspection of the row of reflections indicated provides evidence that this monoclinic thaumatin crystal is in fact twinned. In (b), the reflections from a Bence-Jones protein crystal fall not on a single reciprocal lattice but multiple, interwoven lattices indicative of twinned or multiple crystals. In (c), a tetragonal crystal of Bence-Jones protein is seriously disordered as evidenced by the smeared, highly mosaic reflections and high background scatter. 

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