Overexpression, purification, and crystal structure of native ER alpha LBD

Several crystal structures of human estrogen receptor alpha ligand-binding domain (hER alpha LBD) complexed with agonist or antagonist molecules have previously been solved. The proteins had been modified in cysteine residues (carboxymethylation) or renatured in urea to circumvent aggregation and de naturation problems. In this work, high-level protein expression and purifi cation together with crystallization screening procedure yielded high amoun ts of soluble protein without renaturation or modifications steps, The nati ve protein crystallizes in the space group P3(2)21 with three molecules in the asymmetric unit. The overall structure is very similar to that previous ly reported for the hER alpha LBD with cysteine carboxymethylated residues thus validating the modification approach. The present strategy can be adap ted to other cases where the solubility and the proper folding is a difficu lty. (C) 2001 Academic Press.