Isomorphous Replacement: A Survey of Recent Developments

Abstract

The heavy atom replacement method [1] remains the major tool to phase new macromolecular structures. A derivative is created by adding a heavy atom compound to the macromolecule, while keeping the crystal lattice. This is possible in the case of macromolecular crystals, since they contain solvent channels which allow the diffusion of heavy atom salts. In this case, the electron densities add, and if the reciprocal lattices are sampled at the same points (isomorphous crystals) the following relation is valid:

$$ Fph = Fp + Fh $$

((1))

where Fp=Fp. exp (iαp) is the macromolecular structure factor, Fh= Fh. exp (iαh) is the heavy atom structure factor and Fph=Fph. exp (iαph) is the derivative structure factor. To apply the method, Fp and Fph are measured experimentally. A Patterson map is then calculated with the coefficients |Fph-Fp|; it shows the peaks corresponding to the heavy atoms plus noise. These peaks are readily identifiable, provided their signal is significantly higher than the noise. They lead to the calculation of the heavy atom positions and, after the refinement of the heavy atom parameters, to the determination of Fh. Equation (1) can then be solved using the amplitudes Fph and Fp. As shown in Figure 1, there are two solutions, symmetrical around Fh, which cause a two fold ambiguity for αp (native phase). This ambiguity can be solved by a second derivative.