Resolution Dependence of an Ab Initio Phasing Method in Protein X-ray Crystallography

Abstract

For direct phasing of protein crystals, a method based on the hybrid-input-output (HIO) algorithm has been proposed and tested on a variety of structures. So far, however, the diffraction data have been limited to high-resolution ones, i.e., higher than 2 Å. In principle, the methodology can be applied to data of lower resolutions, which might be particularly useful for phasing membrane protein crystals. For resolutions higher than 3.5 Å, it seems the atomic structure is solvable. For data of lower resolutions, information of the secondary structures and the protein boundary can still be obtained. Examples are given to support the conclusions. Real experimental data are used. Two aspects of the observed data have been discussed: removal of the measured low-resolution reflections and involvement of the unmeasured high-resolution reflections. The ab initio phasing employs histogram matching for density modification. A question arises whether the reference histogram used should match the resolution of the diffraction data or not. It seems that there is an optimal histogram which is good to use for data at various resolutions.

Direct Phasing of Protein Crystals with Non-Crystallographic Symmetry

“…Direct phasing of protein crystals has been developed in our previous articles [12][13][14][15]. It divides a unit cell into a grid and starts from random densities in an asymmetric unit (ASU).…”

“…σ 2 is chosen to be 5.0 Å throughout the iterations. σ 3 is chosen to be 4 Å at the first iteration, decreasing linearly in the following iterations [12,15]. At the last one thousand iterations, σ 3 is reduced to 2.5 Å, and it stays constant when solvent flattening [32] is applied during the last one thousand iterations.…”

“…The success rate was about 25%. The final mean phase error of a successful run was about 49 • which could be further reduced to 39 • by averaging over the successful runs [15] as shown in Figure 6. The final density is good enough for automatic model building by ARP/wARP [34].…”

“…Iterative phasing algorithms have been proposed to solve the phase problem of protein crystallography [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15]. Liu et al showed that direct phasing of protein structures is possible when the protein boundary is known or can be derived from other experiments, such as small-angle X-ray scattering [8].…”

Trial Direct Phasing Calculation of A Thyroid Hormone Receptor Alpha Structure (4LNW)

“…The 200 runs clearly fall into two clusters on the diagram, with the successful runs corresponding to a smaller R f ree and R work . A close look at the density maps of the successful runs reveals small differences; therefore, 30 successful runs with the lowest Rs were averaged over [6] for model building with the CCP4 [7] software. The reconstructed model and the calculated protein boundary are shown in Figure 3.…”