Kinetic and mutational studies of three NifS homologs from Escherichia coli: mechanistic difference between L-cysteine desulfurase and L-selenocysteine lyase reactions.

  title={Kinetic and mutational studies of three NifS homologs from Escherichia coli: mechanistic difference between L-cysteine desulfurase and L-selenocysteine lyase reactions.},
  author={H. Mihara and T. Kurihara and T. Yoshimura and N. Esaki},
  journal={Journal of biochemistry},
  volume={127 4},
We have purified three NifS homologs from Escherichia coli, CSD, CsdB, and IscS, that appear to be involved in iron-sulfur cluster formation and/or the biosynthesis of selenophosphate. All three homologs catalyze the elimination of Se and S from L-selenocysteine and L-cysteine, respectively, to form L-alanine. These pyridoxal 5'-phosphate enzymes were inactivated by abortive transamination, yielding pyruvate and a pyridoxamine 5'-phosphate form of the enzyme. The enzymes showed non-Michaelis… Expand
Escherichia coli NifS-like Proteins Provide Selenium in the Pathway for the Biosynthesis of Selenophosphate*
The ability of the NifS-like proteins to function as a selenium delivery protein for the in vitro biosynthesis of selenophosphate by E. coli wild-type SPS and the activation of the SPS (C17S) mutant suggest a seenium delivery function for the proteins in vivo. Expand
Reaction Mechanism and Molecular Basis for Selenium/Sulfur Discrimination of Selenocysteine Lyase*
Findings provide, for the first time, the basis for understanding how trace amounts of a selenium-containing substrate is distinguished from excessive amounts of its cognate sulfur-containing compound in a biological system. Expand
The iscS gene is essential for the biosynthesis of 2-selenouridine in tRNA and the selenocysteine-containing formate dehydrogenase H
  • H. Mihara, S. Kato, +6 authors N. Esaki
  • Chemistry, Medicine
  • Proceedings of the National Academy of Sciences of the United States of America
  • 2002
Evidence is reported that a strain lacking IscS is incapable of synthesizing 5-methylaminomethyl-2-selenouridine and its precursor 5- methylamine-3-thiouridine in tRNA, suggesting that the sulfur atom released from l-cysteine by the action of IscC is incorporated into mnm5s2U. Expand
A Sulfurtransferase Is Required in the Transfer of Cysteine Sulfur in the in Vitro Synthesis of Molybdopterin from Precursor Z in Escherichia coli *
It was found that MPT can be produced from precursor Z in an E. coli iscS mutant strain, indicating that IscS is not required for the in vivo sulfuration of MPT synthase, and a comparison of the ability of the three sulfurtransferases to provide the sulfur for MPT formation showed the highest activity for CSD in the in vitro system. Expand
Examining the function of the residue C 207 in the cysteine desulfurase reaction of an essential NifS-like protein from Synechocystis PCC 6803
Biochemical analysis showed the NifS protein is essential to the production of many metalloenzymes, specifically iron-sulfur clusters. Slr0077 is thought to play a vital role in the shuttling ofExpand
Requirement for IscS in Biosynthesis of All Thionucleosides in Escherichia coli
It was found that unfractionated tRNA isolated from the iscS mutant strain contained <5% of the level of sulfur found in the parent strain, and an overall scheme for thionucleoside biosynthesis in E. coli is discussed. Expand
Gene Cloning, Purification, and Characterization of Two Cyanobacterial NifS Homologs Driving Iron-Sulfur Cluster Formation
The results suggest that SsCsd1 and SSCsd2 facilitate the iron-sulfur cluster assembly by producing inorganic sulfur from L-cysteine, which probably proceeds through a mechanism that is different from that in A. vinelandii. Expand
Selenium Is Mobilized In Vivo from Free Selenocysteine and Is Incorporated Specifically into Formate Dehydrogenase H and tRNA Nucleosides
During growth of E. coli in the presence of 0.1 microM (75)SeO(3)(2-) and increasing amounts of L-selenocysteine, a concomitant decrease in formate dehydrogenase H and nucleosides of bulk tRNA was observed, consistent with the mobilization of selenium from L-selsenocy steine in vivo and its use in selenophosphate formation. Expand
Gene Cloning , Purification , and Characterization of Two Cy Homologs Driving Iron-Sulfur Cluster Formation
isms. Cysteine desulfurase, originally identified in a nitrogen-fixing bacterium, Azotobacter vinelandii, and named NifS, is essential for the production of a holo-form of nitrogenase in vivo. TheExpand
Substitutions in an Active Site Loop of Escherichia coli IscS Result in Specific Defects in Fe-S Cluster and Thionucleoside Biosynthesis in Vivo*
The results indicate that residues in the active site loop can selectively affect Fe-S cluster biosynthesis in vivo without detectably affecting persulfide delivery and suggest that additional assays may be necessary to fully represent the functions of IscS in Fe- S cluster formation. Expand