Kinetic and thermodynamic properties of two barley thioredoxin h isozymes, HvTrxh1 and HvTrxh2

Kenji Maeda; Per Hägglund; Olof Björnberg; Jakob R Winther; Birte Svensson

doi:10.1016/j.febslet.2010.06.028

Kinetic and thermodynamic properties of two barley thioredoxin h isozymes, HvTrxh1 and HvTrxh2

Kenji Maeda, Per Hägglund, Olof Björnberg, Jakob R Winther, Birte Svensson

Biomolecular Sciences

15 Citations (Scopus)

Abstract

Barley thioredoxin h isozymes 1 (HvTrxh1) and barley thioredoxin h isozymes 2 (HvTrxh2) show distinct spatiotemporal distribution in germinating seeds. Using a novel approach involving measurement of bidirectional electron transfer rates between Escherichia coli thioredoxin, which exhibits redox-dependent fluorescence, and the barley isozymes, reaction kinetics and thermodynamic properties were readily determined. The reaction constants were ∼60% higher for HvTrxh1 than HvTrxh2, while their redox potentials were very similar. The primary nucleophile, Cys_N, of the active site Trp-Cys_N-Gly-Pro-Cys_C motif has an apparent pK_a of 7.6 in both isozymes, as found by iodoacetamide titration, but showed ∼70% higher reactivity in HvTrxh1, suggesting significant functional difference between the isozymes.

Original language	English
Journal	FEBS Letters
Volume	584
Issue number	15
Pages (from-to)	3376-3380
Number of pages	4
ISSN	0014-5793
DOIs	https://doi.org/10.1016/j.febslet.2010.06.028
Publication status	Published - Aug 2010

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@article{b86d2db0a15111df928f000ea68e967b,

title = "Kinetic and thermodynamic properties of two barley thioredoxin h isozymes, HvTrxh1 and HvTrxh2",

abstract = "Barley thioredoxin h isozymes 1 (HvTrxh1) and barley thioredoxin h isozymes 2 (HvTrxh2) show distinct spatiotemporal distribution in germinating seeds. Using a novel approach involving measurement of bidirectional electron transfer rates between Escherichia coli thioredoxin, which exhibits redox-dependent fluorescence, and the barley isozymes, reaction kinetics and thermodynamic properties were readily determined. The reaction constants were ∼60% higher for HvTrxh1 than HvTrxh2, while their redox potentials were very similar. The primary nucleophile, CysN, of the active site Trp-CysN-Gly-Pro-CysC motif has an apparent pKa of 7.6 in both isozymes, as found by iodoacetamide titration, but showed ∼70% higher reactivity in HvTrxh1, suggesting significant functional difference between the isozymes.",

author = "Kenji Maeda and Per H{\"a}gglund and Olof Bj{\"o}rnberg and Winther, {Jakob R} and Birte Svensson",

note = "Keywords: Thioredoxin; Dithiol/disulfide exchange; Tryptophan fluorescence; Redox potential; Thiol pKa",

year = "2010",

month = aug,

doi = "10.1016/j.febslet.2010.06.028",

language = "English",

volume = "584",

pages = "3376--3380",

journal = "F E B S Letters",

issn = "0014-5793",

publisher = "JohnWiley & Sons Ltd",

number = "15",

}

TY - JOUR

T1 - Kinetic and thermodynamic properties of two barley thioredoxin h isozymes, HvTrxh1 and HvTrxh2

AU - Maeda, Kenji

AU - Hägglund, Per

AU - Björnberg, Olof

AU - Winther, Jakob R

AU - Svensson, Birte

N1 - Keywords: Thioredoxin; Dithiol/disulfide exchange; Tryptophan fluorescence; Redox potential; Thiol pKa

PY - 2010/8

Y1 - 2010/8

N2 - Barley thioredoxin h isozymes 1 (HvTrxh1) and barley thioredoxin h isozymes 2 (HvTrxh2) show distinct spatiotemporal distribution in germinating seeds. Using a novel approach involving measurement of bidirectional electron transfer rates between Escherichia coli thioredoxin, which exhibits redox-dependent fluorescence, and the barley isozymes, reaction kinetics and thermodynamic properties were readily determined. The reaction constants were ∼60% higher for HvTrxh1 than HvTrxh2, while their redox potentials were very similar. The primary nucleophile, CysN, of the active site Trp-CysN-Gly-Pro-CysC motif has an apparent pKa of 7.6 in both isozymes, as found by iodoacetamide titration, but showed ∼70% higher reactivity in HvTrxh1, suggesting significant functional difference between the isozymes.

AB - Barley thioredoxin h isozymes 1 (HvTrxh1) and barley thioredoxin h isozymes 2 (HvTrxh2) show distinct spatiotemporal distribution in germinating seeds. Using a novel approach involving measurement of bidirectional electron transfer rates between Escherichia coli thioredoxin, which exhibits redox-dependent fluorescence, and the barley isozymes, reaction kinetics and thermodynamic properties were readily determined. The reaction constants were ∼60% higher for HvTrxh1 than HvTrxh2, while their redox potentials were very similar. The primary nucleophile, CysN, of the active site Trp-CysN-Gly-Pro-CysC motif has an apparent pKa of 7.6 in both isozymes, as found by iodoacetamide titration, but showed ∼70% higher reactivity in HvTrxh1, suggesting significant functional difference between the isozymes.

U2 - 10.1016/j.febslet.2010.06.028

DO - 10.1016/j.febslet.2010.06.028

M3 - Journal article

C2 - 20594550

SN - 0014-5793

VL - 584

SP - 3376

EP - 3380

JO - F E B S Letters

JF - F E B S Letters

IS - 15

ER -

Kinetic and thermodynamic properties of two barley thioredoxin h isozymes, HvTrxh1 and HvTrxh2

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