Open AccessStructures of human thymidylate synthase R163K with dUMP, FdUMP and glutathione show asymmetric ligand binding
Author(s) -
Gibson Lydia M.,
Celeste Lesa R.,
Lovelace Leslie L.,
Lebioda Lukasz
Publication year - 2011
Publication title -
acta crystallographica section d
Language(s) - English
Resource type - Journals
ISSN - 1399-0047
DOI - 10.1107/s0907444910044732
Subject(s) - dimer , glutathione , thymidylate synthase , covalent bond , chemistry , ligand (biochemistry) , adduct , active site , protein subunit , stereochemistry , nucleophile , enzyme , biochemistry , catalysis , biology , organic chemistry , receptor , cancer , fluorouracil , gene , genetics
Thymidylate synthase (TS) is a well validated target in cancer chemotherapy. Here, a new crystal form of the R163K variant of human TS (hTS) with five subunits per asymmetric part of the unit cell, all with loop 181–197 in the active conformation, is reported. This form allows binding studies by soaking crystals in artificial mother liquors containing ligands that bind in the active site. Using this approach, crystal structures of hTS complexes with FdUMP and dUMP were obtained, indicating that this form should facilitate high‐throughput analysis of hTS complexes with drug candidates. Crystal soaking experiments using oxidized glutathione revealed that hTS binds this ligand. Interestingly, the two types of binding observed are both asymmetric. In one subunit of the physiological dimer covalent modification of the catalytic nucleophile Cys195 takes place, while in another dimer a noncovalent adduct with reduced glutathione is formed in one of the active sites.