Published in
Nature Research, Nature Structural and Molecular Biology, 8(16), p. 890-896, 2009
DOI: 10.1038/nsmb.1627
Nature Research, Nature Structural and Molecular Biology, 12(16), p. 1331-1331, 2009
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Diversity of Chemical Mechanisms in Thioredoxin Catalysis Revealed by Single-Molecule Force Spectroscopy
,
Pallav Kosuri,
Inmaculada Sanchez-Romero,
Arun P. Wiita,
David Rodriguez-Larrea,
Ana Chueca,
Arne Holmgren,
Antonio Miranda-Vizuete,
Katja Becker,
Seung-Hyun Cho,
Jon Beckwith,
Eric Gelhaye,
Jean P. Jacquot,
Eric A. Gaucher
and other authors.
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Abstract
Thioredoxins (Trxs) are oxidoreductase enzymes, present in all organisms, that catalyze the reduction of disulfide bonds in proteins. By applying a calibrated force to a substrate disulfide, the chemical mechanisms of Trx catalysis can be examined in detail at the single-molecule level. Here we use single-molecule force-clamp spectroscopy to explore the chemical evolution of Trx catalysis by probing the chemistry of eight different Trx enzymes. All Trxs show a characteristic Michaelis-Menten mechanism that is detected when the disulfide bond is stretched at low forces, but at high forces, two different chemical behaviors distinguish bacterial-origin from eukaryotic-origin Trxs. Eukaryotic-origin Trxs reduce disulfide bonds through a single-electron transfer reaction (SET), whereas bacterial-origin Trxs show both nucleophilic substitution (S(N)2) and SET reactions. A computational analysis of Trx structures identifies the evolution of the binding groove as an important factor controlling the chemistry of Trx catalysis.