6o12 | pdb_00006o12
E. coli cysteine desulfurase SufS H123A
Structural highlights
FunctionSUFS_ECOLI Cysteine desulfurases mobilize the sulfur from L-cysteine to yield L-alanine, an essential step in sulfur metabolism for biosynthesis of a variety of sulfur-containing biomolecules. Component of the suf operon, which is activated and required under specific conditions such as oxidative stress and iron limitation. Acts as a potent selenocysteine lyase in vitro, that mobilizes selenium from L-selenocysteine. Selenocysteine lyase activity is however unsure in vivo.[1] [2] [3] [4] [5] Publication Abstract from PubMedIron-sulfur (Fe-S) clusters are necessary for the proper functioning of numerous metalloproteins. Fe-S cluster (Isc) and sulfur utilization factor (Suf) pathways are the key biosynthetic routes responsible for generating these Fe-S cluster prosthetic groups in Escherichia coli Whereas Isc dominates under normal conditions, Suf takes over during periods of iron depletion and oxidative stress. Sulfur acquisition via these systems relies on the ability to remove sulfur from free cysteine using a cysteine desulfurase mechanism. In the Suf pathway, the dimeric SufS protein uses the cofactor pyridoxal-5'-phosphate (PLP) to abstract sulfur from free cysteine, resulting in the production of alanine and persulfide. Despite much progress, the stepwise mechanism by which this PLP-dependent enzyme operates remains unclear. Here, using rapid-mixing kinetics in conjunction with X-ray crystallography, we analyzed the pre-steady state kinetics of this process while assigning early intermediates of the mechanism. We employed H123A and C364A SufS variants to trap Cys-aldimine and Cys-ketimine intermediates of the cysteine desulfurase reaction, enabling direct observations of these intermediates and associated conformational changes of the SufS active site. Of note, we propose that Cys-364 is essential for positioning the Cys-aldimine for Calpha deprotonation, His-123 acts to protonate the Ala-eneamine intermediate, and Arg-56 facilitates catalysis by hydrogen bonding with the sulfhydryl of Cys-aldimine. Our results, along with previous SufS structural findings, suggest a detailed model of the SufS-catalyzed reaction from Cys binding to C-S bond formation and indicate that Arg-56, His-123, and Cys-364 are critical SufS residues in this C-S bond cleavage pathway. Direct observation of intermediates in the SufS cysteine desulfurase reaction reveals functional roles of conserved active-site residues.,Blahut M, Wise CE, Bruno MR, Dong G, Makris TM, Frantom PA, Dunkle JA, Outten FW J Biol Chem. 2019 Jun 27. pii: RA119.009471. doi: 10.1074/jbc.RA119.009471. PMID:31248989[6] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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