Evidence for cysteine persulfide as reaction product of L-Cyst(e)ine C-S-Lyase (C-DES) from Synechocystis

The pyridoxal phosphate-dependent monomeric L-cysteine/cystine C-S-lyase (C-DES), previously isolated from Synechocystis PCC 6714 by its capacity to direct [2Fe-2S] cluster assembly of ferredoxin in vitro (Leibrecht, I., and Kessler, D, (1997) J, Biol, Chem, 272, 10442-10447), has now been cloned, sequenced, and overexpressed in Escherichia cell. The amino acid sequence of C-DES was found to be nearly identical (92% identity) to the open reading frame slr2143 of Synechocystis PCC 6803 and showed a more distant relationship to the NifS family of proteins (about 27% identity). Recombinant C-DES displayed activities equal to the isolate from Synechocystis in terms of the cyst(e)ine lyase reaction and holoferre-doxin formation which recommended its use for functional and mechanistic studies. Investigation of the substrate spectrum for beta-elimination found L-cysteine to be a poor substrate (k(cat) approximate to 0.15 s(-1)) in contrast to L-cystine (k(cat) = 36 s(-1)) and several related compounds. Of these compounds, desaminocystine (S-(carboxyethylthio)-L-cysteine) was used for C-DES-mediated persulfide generation. Stabilization of the linear persulfide 3-(disulfanyl)-propionic acid was achieved by cyclization as a novel intramolecular trapping reaction; this yielded 1,2-dithiolan-3-one which was isolated and identified by chemical analyses.