Initiating heavy-atom-based phasing by multi-dimensional molecular replacement
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To obtain an electron-density map from a macromolecular crystal the phase problem needs to be solved, which often involves the use of heavy-atom derivative crystals and concomitant heavy-atom substructure determination. This is typically performed by dual-space methods, direct methods or Patterson-based approaches, which however may fail when only poorly diffracting derivative crystals are available. This is often the case for, for example, membrane proteins. Here, an approach for heavy-atom site identification based on a molecular-replacement parameter matrix (MRPM) is presented. It involves an n-dimensional search to test a wide spectrum of molecular-replacement parameters, such as different data sets and search models with different conformations. Results are scored by the ability to identify heavy-atom positions from anomalous difference Fourier maps. The strategy was successfully applied in the determination of a membrane-protein structure, the copper-transporting P-type ATPase CopA, when other methods had failed to determine the heavy-atom substructure. MRPM is well suited to proteins undergoing large conformational changes where multiple search models should be considered, and it enables the identification of weak but correct molecular-replacement solutions with maximum contrast to prime experimental phasing efforts.
Originalsprog | Engelsk |
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Tidsskrift | Acta crystallographica Section D: Structural biology |
Vol/bind | 72 |
Udgave nummer | 3 |
Sider (fra-til) | 440-445 |
Antal sider | 6 |
ISSN | 2059-7983 |
DOI | |
Status | Udgivet - 1 mar. 2016 |
Eksternt udgivet | Ja |
ID: 157959724