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

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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 approximately 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 approximately 70% higher reactivity in HvTrxh1, suggesting significant functional difference between the isozymes.
Original languageEnglish
JournalFEBS Letters
Issue number15
Pages (from-to)3376-3380
Number of pages4
Publication statusPublished - 2010

Bibliographical note

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

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