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Influence of the p K a value of the buried, active‐site cysteine on the redox properties of thioredoxin‐like oxidoreductases
Author(s) -
Mössner Ekkehard,
Iwai Hideo,
Glockshuber Rudi
Publication year - 2000
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(00)01738-5
Subject(s) - cysteine , thioredoxin , chemistry , active site , thiol , oxidizing agent , redox , ferredoxin thioredoxin reductase , stereochemistry , nucleophile , enzyme , biochemistry , catalysis , thioredoxin reductase , organic chemistry
Thioredoxin constitutes the prototype of the thiol‐disulfide oxidoreductase family. These enzymes contain an active‐site disulfide bridge with the consensus sequence Cys‐Xaa‐Xaa‐Cys. The more N‐terminal active‐site cysteine is generally a strong nucleophile with an abnormal low p K a value. In contrast, the more C‐terminal cysteine is buried and only little is known about its effective p K a during catalysis of disulfide exchange reactions. Here we have analyzed the p K a values of the active‐site thiols in wild type thioredoxin and a 400‐fold more oxidizing thioredoxin variant by NMR spectroscopy, using selectively 13 C β ‐Cys‐labeled proteins. We find that the effective p K a of the buried cysteine (p K b ) of the variant is increased, while the p K a of the more N‐terminal cysteine (p K N ) is decreased relative to the corresponding p K a values in the wild type. We propose two empirical models which exclusively require the knowledge of p K N to predict the redox properties of thiol‐disulfide oxidoreductases with reasonable accuracy.