Literature summary for 2.5.1.73 extracted from

Fukunaga, R.; Yokoyama, S.
Structural insights into the second step of RNA-dependent cysteine biosynthesis in archaea: crystal structure of Sep-tRNA:Cys-tRNA synthase from Archaeoglobus fulgidus (2007), J. Mol. Biol., 370, 128-141.

Cloned(Commentary)

Cloned (Comment)	Organism
SepCysS1, overexpression in Escherichia coli strain BL21(DE3), expression of selenomethionine-labeled SepCysS1 in Escherichia coli strain B834	Archaeoglobus fulgidus

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant wild-type SepVysS1 and selenomethionine-labeled SepCysS1, hanging-drop vapor diffusion method, 0.001 ml protein solution is mixed with 0.001 ml reservoir solution containing 80 mM sodium acetate buffer, pH 4.4, 160 mM NaCl, and 1.00 M ammonium sulfate, 20°C, equilibration against 0.5 ml reservoir solution, cryoprotection by 25% v/v glycerol, X-ray diffraction structure determination and analysis at 2.4-3.2 A resolution, modeling	Archaeoglobus fulgidus

Crystallization (Comment)

Organism

purified recombinant wild-type SepVysS1 and selenomethionine-labeled SepCysS1, hanging-drop vapor diffusion method, 0.001 ml protein solution is mixed with 0.001 ml reservoir solution containing 80 mM sodium acetate buffer, pH 4.4, 160 mM NaCl, and 1.00 M ammonium sulfate, 20°C, equilibration against 0.5 ml reservoir solution, cryoprotection by 25% v/v glycerol, X-ray diffraction structure determination and analysis at 2.4-3.2 A resolution, modeling

Archaeoglobus fulgidus

Metals/Ions

Metals/Ions	Comment	Organism	Structure
sulfate	is bound in the proximity of PLP by the side-chains of the conserved Arg79, His103, and Tyr104 residues, the PLP-bound active site is located deep within the large, basic cleft for recognizing Sep-tRNACys	Archaeoglobus fulgidus

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
additional information	Archaeoglobus fulgidus	two-step Cys-tRNACys formation: in organisms like Archaeoglobus fulgidus lacking a canonical cysteinyl-tRNA synthetase for the direct Cys-tRNACys formation, Cys-tRNACys is produced by the indirect pathway, in which the non-canonical O-phosphoseryl-tRNA synthetase, SepRS, ligates the non-canonical amino acid O-phosphoserine, Sep, to tRNACys, and the Sep-tRNA:Cys-tRNA synthase converts the produced Sep-tRNACys to Cys-tRNACys, overview, the SepRS/SepCysS pathway is the sole route for cysteine biosynthesis in the organism	?	-	?
O-phospho-L-seryl-tRNACys + sulfate	Archaeoglobus fulgidus	the in vivo sulfur donor is not determined	L-cysteinyl-tRNACys + phosphate	-	?

Organism

Organism	UniProt	Comment	Textmining
Archaeoglobus fulgidus	O30207	SepCysS1; SepCysS1	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant SepCysS1 from Escherichia coli strain BL21(DE3) by anion exchange chromatography and affinity chromatography on a heparin resin, recombinant selenomethionine-labeled SepCysS1 from Escherichia coli strain B834	Archaeoglobus fulgidus

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
additional information	two-step Cys-tRNACys formation: in organisms like Archaeoglobus fulgidus lacking a canonical cysteinyl-tRNA synthetase for the direct Cys-tRNACys formation, Cys-tRNACys is produced by the indirect pathway, in which the non-canonical O-phosphoseryl-tRNA synthetase, SepRS, ligates the non-canonical amino acid O-phosphoserine, Sep, to tRNACys, and the Sep-tRNA:Cys-tRNA synthase converts the produced Sep-tRNACys to Cys-tRNACys, overview, the SepRS/SepCysS pathway is the sole route for cysteine biosynthesis in the organism	Archaeoglobus fulgidus	?	-	?
additional information	the active site contains an internal aldimine Lys209-PLP and the sulfate ion, SepCysS should not bind Sep-tRNASec and discriminate tRNACys from tRNASec on the basis of the differences in the length of the T-arms, or SepCysS recognizes the discriminator sequence, which is Ura73 in tRNACys and Gua73 in tRNASec, overview	Archaeoglobus fulgidus	?	-	?
O-phospho-L-seryl-tRNACys + sulfate	the in vivo sulfur donor is not determined	Archaeoglobus fulgidus	L-cysteinyl-tRNACys + phosphate	-	?
O-phospho-L-seryl-tRNACys + sulfide	modeling of tRNA binding, overview, sulfide, persulfide, and thiosulfate, but not cysteine, can function as sulfur donor in vitro, the active site is located deep within the large, basic cleft to accommodate Sep-tRNACys, binding modeling of Sep-tRNACys, overview, possibly the side-chain of a Cys residue in SepCysS becomes persulfided as a sulfur transfer intermediate state	Archaeoglobus fulgidus	L-cysteinyl-tRNACys + phosphate	-	?

Subunits

Subunits	Comment	Organism
dimer	the active site is located near the dimer interface, crystal structure analysis, overview	Archaeoglobus fulgidus
More	amino acid residue conservation mapping on the basis of the surface electrostatic potential, overview, construction of a SepRS-tRNACys-SepCysS ternary complex model, in the ternary complex the phosphoserylated 3'-terminus of tRNACys can possibly be transferred directly from SepRS to SepCysS, for conversion to the cysteinylated form, overview	Archaeoglobus fulgidus

Synonyms

Synonyms	Comment	Organism
SepCysS	-	Archaeoglobus fulgidus

Cofactor

Cofactor	Comment	Organism	Structure
pyridoxal 5'-phosphate	dependent on, pyridoxal 5'-phosphate is covalently bound to the side-chain of the conserved Lys209 at the active site	Archaeoglobus fulgidus