Leslie B. Poole, PhD, Wake Forest University — Published: November 5, 2012
The need for chemical probes to detect and identify oxidized cysteine residues in proteins, particularly in their sulfenic acid state, became clear to me as a graduate student in the 1980’s while working on the cysteine-dependent peroxide-reducing enzyme from lactococci, NADH peroxidase. The natural oxidized form of NADH peroxidase contains cysteine sulfenic acid (plus an oxidized, noncovalently bound FAD), but this was not so straightforward to prove (Poole & Claiborne, 1989, J Biol Chem 264, 12330).
Later, working with bacterial peroxiredoxins, we pioneered the use of NBD chloride as a trapping agent capable of discriminating between thiol and sulfenic acid moieties on proteins (Ellis & Poole, 1997, Biochemistry 36, 15013), but the utility of this approach was largely restricted to purified proteins with only one or a small number of cysteines.
Wishing to test the hypothesis that sulfenic acid formation plays a much broader role in biological systems than was yet appreciated, I applied for and obtained support from an Established Investigatorship from the AHA to follow my dream of coming up with a reagent to tag sulfenic acids in proteins using a dimedone-like reagent linked to a reporter group. Bruce King in Chemistry at Wake Forest was very interested in this project. He helped develop the first few reagents for testing in our biochemical assays (DCP-Bio1 and DCP-N3). These reagents formed the basis of our patents issued over the next few years (in 2007 and 2010).
Thus, starting with a proof-of-principle experiment conducted by a summer undergraduate researcher, the synthesis of these and new generations of reagents, including those created by colleague Cristina Furdui (Biotin-1,3-cyclopentanedione (BP1), has greatly expanded the arsenal of reagents available to us in exploring the largely unmapped territory of protein oxidation chemistry within living cells. This has enabled a new era of exploration for our lab and others into the mechanisms underlying redox responses in cells.
Cysteine sulfenic acid, protein oxidation, DCP-Bio1, DCP-N3, Biotin-1,3-cyclopentanedione (BP1), Free Methionine-(R)-Sulfoxide Reductase (C84S, C94S)