Direct phasing of protein crystals with high solvent content

Abstract

An iterative transform method is proposed for solving the phase problem in protein crystallography. In each iteration, a weighted average electron-density map is constructed to define an estimated protein mask. Solvent flattening is then imposed through the hybrid input-output algorithm [Fienup (1982). Appl. Opt. 21, 2758-2769]. Starting from random initial phases, after thousands of iterations the mask evolves into the correct shape and the phases converge to the correct values with an average error of 30-40° for high-resolution data for several protein crystals with high solvent content. With the use of non-crystallographic symmetry, the method could potentially be extended to phase protein crystals with less than 50% solvent fraction. The new phasing algorithm can supplement and enhance the traditional refinement tools.

Improving the efficiency of molecular replacement by utilizing a new iterative transform phasing algorithm

“…The radius of the weighting function started from a medium value such as 3 Å at the beginning of the iteration and decreased slowly to a smaller value, for example, 2 Å , at the end of the iteration. Such values are smaller than the ones used before in ab initio calculations (He & Su, 2015), because the placed template provides significantly better starting density.…”

“…A weighted-average electron-density map was calculated to identify a protein mask from the calculated electron-density map. The HIO method has proved to be a very effective solvent-flattening and phaserecovery technique (Liu et al, 2012;He & Su, 2015;Ayyer et al, 2016) by consistently applying constraints in real space and Fourier space (Marchesini, 2007). Like real-space phasing methods (Su, 2008), the protein mask serves as a high-density support.…”

“…The solvent content of 3tx8 is large enough for ab initio phasing (He & Su, 2015). Starting from random phases, we got three successful runs among 15 attempts.…”

“…During the iterations, the radius of the weighting function was varied from 4 to 2 Å . The choice of a larger value of (He & Su, 2015) would result in a slightly larger phase error. Fig.…”

“…There are several other approaches to overcome this problem (Schwarzenbacher et al, 2004;Keller et al, 2006;He et al, 2007;Bunkó czi & Read, 2011;Brunger et al, 2012;Sammito et al, 2014;Millá n et al, 2015;Carrozzini et al, 2015). In this paper we consider an alternate improvement of MR based on an extension of the hybrid input-output (HIO) phasing algorithm (Fienup, 1982;He & Su, 2015). The new phasing scheme can be regarded as a powerful extension of the conventional solvent-flattening-related density modification.…”