NMR structure of oxidized glutaredoxin 3 from Escherichia coli

A high precision NMR structure of oxidized glutaredoxin 3 [C65Y] from Esche richia coli has been determined. The conformation of the active site includ ing the disulphide bridge is highly similar to those in glutaredoxins from pig liver and T4 phage. A comparison with the previously determined structu re of glutaredoxin 3 [C14S, C65Y] in a complex with glutathione reveals con formational changes between the free and substrate-bound form which include s the sidechain of the conserved, active site tyrosine residue. In the oxid ized form this tyrosine is solvent exposed, while it adopts less exposed co nformation, stabilized by hydrogen bonds, in the mixed disulfide with gluta thione. The structures further suggest that the formation of a covalent lin kage between glutathione and glutaredoxin 3 is necessary in order to induce these structural changes upon binding of the glutathione peptide. This cou ld explain the observed low affinity of glutaredoxins for S-blocked glutath ione analogues, in spite of the fact that glutaredoxins are highly specific reductants of glutathione mixed disulfides. (C) 2000 Academic Press.