Expression, purification, crystallization and preliminary crystallographic analysis of the mouse transcription factor MafB in complex with its DNA‐recognition motif Cmare
The MafB transcription factor (residues 211–305) has been overexpressed in and purified from Escherichia coli. A protein–DNA complex between the MafB homodimer and the 21 bp Maf‐recognition sequence known as Cmare has been successfully reconstituted in vitro and subsequently crystallized. The diffraction properties of the protein–DNA complex crystals were improved using a combination of protein‐construct boundary optimization and targeted mutagenesis to promote crystal lattice stability. Both native and mercury‐derivatized crystals have been prepared using these optimized conditions. The crystals belong to space group P41212 or P43212, with unit‐cell parameters a = b = 94.8, c = 197.9 Å. An anomalous difference Patterson map computed using data collected from crystals grown in the presence of HgCl2 reveals four peaks. This corresponds to two copies of the protein–DNA complex in the asymmetric unit, with a solvent content of 62% and a Matthews coefficient of 3.22 Å3 Da−1.
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