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Direct NMR observation of the Cys‐14 thiol proton of reduced Escherichia coli glutaredoxin‐3 supports the presence of an active site thiol‐thiolate hydrogen bond
Author(s) -
Nordstrand Kerstin,
Åslund Fredrik,
Meunier Sylvie,
Holmgren Arne,
Otting Gottfried,
Berndt Kurt D
Publication year - 1999
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(99)00401-9
Subject(s) - glutaredoxin , thiol , cysteine , chemistry , active site , hydrogen bond , proton nmr , escherichia coli , proton , resonance (particle physics) , crystallography , stereochemistry , biochemistry , enzyme , glutathione , molecule , organic chemistry , physics , particle physics , quantum mechanics , gene
The active site of Escherichia coli glutaredoxin‐3 (Grx3) consists of two redox active cysteine residues in the sequence ‐C 11 ‐P‐Y‐C 14 ‐H‐. The 1 H NMR resonance of the cysteine thiol proton of Cys‐14 in reduced Grx3 is observed at 7.6 ppm. The large downfield shift and NOEs observed with this thiol proton resonance suggest the presence of a hydrogen bond with the Cys‐11 thiolate, which is shown to have an abnormally low p K a value. A hydrogen bond would also agree with activity data of Grx3 active site mutants. Furthermore, the activity is reduced in a Grx3 H15V mutant, indicating electrostatic contributions to the stabilization of the Cys‐11 thiolate.