Solvent flattening

Often the initial phases from one of the above techniques (particularly MIR and MAD) are not sufficiently accurate to build in all the atoms and further improvement of the phases must be carried out. The standard approach for this is called density modification, or solvent-flattening. The idea is that if a unit cell contains a sufficient amount of solvent in it (usually around 50%, which is reasonable for most protein crystals), you will be able to use this information to improve the phases for the protein. The region of the unit cell that contains the solvent is set to an average value and new structure factors and phases are then calculated for... 

Why does Fourier cycling improve phases in solvent flattening ... 

Now return to Eq. 3, which describes the process of solvent flattening. As a restraint, it can be written down as follows ... 

Figure 10.3 A graphical representation of solvent flattening in real space, (a) represents a one- dimensional solvent mask, (b) is a one-dimensional, unflattened electron density, and (c) is the resulting flattened electron density map that imposes the solvent mask.

A graphical representation of solvent flattening in real space is shown in Fig. 10.3. In Eq. 8, we multiply the two functions Pinit(x) and g(x) as we flatten the density within the solvent region. However, multiplication in real space is equivalent to a convolution in reciprocal space. Therefore, we can rewrite Eq. 8 as follows ... 

The crude molecular image seen in the F0 map, which is obtained from the original indexed intensity data (IFobsI) and the first phase estimates (a calc), serves now as a model of the desired structure. A crude electron density function is devised to describe the unit-cell contents as well as they can be observed in the first map. Then the function is modified to make it more realistic in the light of known properties of proteins and water in crystals. This process is called, depending on the exact details of procedure, density modification, solvent leveling, or solvent flattening. 

After the calculation of MIR phases, the electron density was improved by solvent flattening followed by non-crystallographic symmetry averaging. The map after solvent flattening showed most of the secondary structure. However, the connectivity of electron density was ambiguous in the loop regions. Since it was clear that the asymmetric unit contained a trimer of GIF related by a non-crystallographic... 

Symmetry averaging and solvent flattening to obtain more interpretable maps. 

Solvent flattening A density modification technique useful when a unit cell contains a high proportion of solvent, as is found in macromolecular structures, which may be 50% or more solvent. An envelope defining the approximate boundary of the molecule is determined from the electron-density map, and all electron density outside this envelope is set to the average value. The usefulness of the method increases as the fraction of solvent in the unit cell increases. 

Usually, isomorphous replacement methods are coupled with a density modification process called solvent flattening. The solvent flattening process was made practical by the introduction of the ISIR/ISAS program suite (Wang, 1985) and other phasing programs such as DM (CCP4,1994) and PHASES (Furey and Swaminathan,... 

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