TY - JOUR
T1 - Initiating heavy-atom-based phasing by multi-dimensional molecular replacement
AU - Pedersen, Bjørn Panyella
AU - Gourdon, Pontus
AU - Liu, Xiangyu
AU - Karlsen, Jesper Lykkegaard
AU - Nissen, Poul
PY - 2016/3/1
Y1 - 2016/3/1
N2 - 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.
AB - 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.
KW - experimental phasing
KW - heavy-atom substructure
KW - molecular replacement
UR - http://www.scopus.com/inward/record.url?scp=85008951539&partnerID=8YFLogxK
U2 - 10.1107/S2059798315022482
DO - 10.1107/S2059798315022482
M3 - Journal article
C2 - 26960131
SN - 2059-7983
VL - 72
SP - 440
EP - 445
JO - Acta crystallographica Section D: Structural biology
JF - Acta crystallographica Section D: Structural biology
IS - Pt 3
ER -