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

Kerstin Nordstrand; Fredrik Åslund; Sylvie Meunier; Arne Holmgren; Gottfried Otting; Kurt D. Berndt

doi:10.1016/S0014-5793(99)00401-9

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

Kerstin Nordstrand, Fredrik Åslund, Sylvie Meunier, Arne Holmgren, Gottfried Otting, Kurt D. Berndt^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

62 Citations (Scopus)

Abstract

The active site of Escherichia coli glutaredoxin-3 (Grx3) consists of two redox active cysteine residues in the sequence -C₁₁-P-Y-C₁₄-H-. The ¹H 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 pK(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. Copyright (C) 1999 Federation of European Biochemical Societies.

Original language	English
Pages (from-to)	196-200
Number of pages	5
Journal	FEBS Letters
Volume	449
Issue number	2-3
DOIs	https://doi.org/10.1016/S0014-5793(99)00401-9
Publication status	Published - 23 Apr 1999
Externally published	Yes

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10.1016/S0014-5793(99)00401-9

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@article{e68f7cd1055b4a6bafe0676abfd4a480,

title = "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",

abstract = "The active site of Escherichia coli glutaredoxin-3 (Grx3) consists of two redox active cysteine residues in the sequence -C11-P-Y-C14-H-. The 1H 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 pK(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. Copyright (C) 1999 Federation of European Biochemical Societies.",

keywords = "Enzyme mechanism, Glutaredoxin, Thiol pK(a)",

author = "Kerstin Nordstrand and Fredrik {\AA}slund and Sylvie Meunier and Arne Holmgren and Gottfried Otting and Berndt, \{Kurt D.\}",

year = "1999",

month = apr,

day = "23",

doi = "10.1016/S0014-5793(99)00401-9",

language = "English",

volume = "449",

pages = "196--200",

journal = "FEBS Letters",

issn = "0014-5793",

publisher = "John Wiley \& Sons Inc.",

number = "2-3",

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TY - JOUR

T1 - 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

AU - Nordstrand, Kerstin

AU - Åslund, Fredrik

AU - Meunier, Sylvie

AU - Holmgren, Arne

AU - Otting, Gottfried

AU - Berndt, Kurt D.

PY - 1999/4/23

Y1 - 1999/4/23

N2 - The active site of Escherichia coli glutaredoxin-3 (Grx3) consists of two redox active cysteine residues in the sequence -C11-P-Y-C14-H-. The 1H 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 pK(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. Copyright (C) 1999 Federation of European Biochemical Societies.

AB - The active site of Escherichia coli glutaredoxin-3 (Grx3) consists of two redox active cysteine residues in the sequence -C11-P-Y-C14-H-. The 1H 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 pK(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. Copyright (C) 1999 Federation of European Biochemical Societies.

KW - Enzyme mechanism

KW - Glutaredoxin

KW - Thiol pK(a)

UR - https://www.scopus.com/pages/publications/0032959801

U2 - 10.1016/S0014-5793(99)00401-9

DO - 10.1016/S0014-5793(99)00401-9

M3 - Article

SN - 0014-5793

VL - 449

SP - 196

EP - 200

JO - FEBS Letters

JF - FEBS Letters

IS - 2-3

ER -

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

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