Information on EC 2.9.1.2 - O-phospho-L-seryl-tRNASec:L-selenocysteinyl-tRNA synthase

O-phospho-L-seryl-tRNASec + selenophosphate

L-selenocysteinyl-tRNASec + phosphate

11 entries

O-phospho-L-seryl-tRNASec + selenophosphate + H2O

L-selenocysteinyl-tRNASec + 2 phosphate

4 entries

O-phospho-L-seryl-tRNASec + selenophosphate + H2O

L-selenocysteinyl-tRNASec + phosphate

additional information

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4 entries

L-phosphoseryl-tRNASec + selenophosphate

L-selenocysteinyl-tRNASec + phosphate

selenocysteine is the only genetically encoded amino acid in humans whose biosynthesis occurs on its cognate transfer RNA (tRNA). O-Phosphoseryl-tRNA:selenocysteinyl-tRNA synthase catalyzes the final step of selenocysteine formation by a tRNA-dependent mechanism

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O-phospho-L-seryl-tRNASec + selenophosphate

L-selenocysteinyl-tRNASec + phosphate

6 entries

O-phospho-L-seryl-tRNASec + selenophosphate + H2O

L-selenocysteinyl-tRNASec + 2 phosphate

4 entries

O-phospho-L-seryl-tRNASec + selenophosphate + H2O

L-selenocysteinyl-tRNASec + phosphate

Methanocaldococcus jannaschii

pyridoxal 5'-phosphate

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Leishmania donovani Bob

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738683

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694847

Q58027

UniProt

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694428, 694847, 723214

UniProt

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primary cultured chicken embryos neurons

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evolution

malfunction

metabolism

physiological function

homotetramer

tetramer

crystal structure of the quaternary complex between human SepSecS, unacylated tRNASec, and a mixture of O-phosphoserine and thiophosphate to 2.8 A resolution

sitting-drop vapor diffusion method at 20°C, crystal structure of the enzyme complexed with pyridoxal 5'-phosphate at 2.5 A resolution

A239T

naturally occurring mutation and site-directed mutagenesis. The mutant forms a stable complex with GroEL. Residue Ala239 is located in helix alpha8 near the site that interacts with the variable arm of tRNASec, and is distant from the active site. The A239T variant binds tRNASec with less affinity compared to wild-type SepSecS, but its catalytic function is unaffected

K173A

in vivo activity of the mutant is indistinguishable from that of the wild-type enzyme

K173M

in vivo activity of the mutant is indistinguishable from that of the wild-type enzyme

Q105A

mutant is inactive in vivo

R313A

mutant is inactive in vivo

R75A

mutant is inactive in vivo

R97A

in vivo activity of the mutant is indistinguishable from that of the wild-type enzyme

R97Q

in vivo activity of the mutant is indistinguishable from that of the wild-type enzyme

T325S

naturally occurring mutation and site-directed mutagenesis, the mutation does not affect the binding affinity of the SepSecS-tRNA complex. The mutant does not form a complex with GroEL. Residue Thr325 is located in helix alpha12 and about 15 A away from the active site. The Thr325 to Ser replacement does not cause any changes in the tetrameric structure of SepSecS. Tetramers of T325S adopt the same structure as wild-type SepSecS. The pathogenic mutation Thr325Ser does not alter the three-dimensional structure of the SepSecS tetramer

Y334C

naturally occurring mutation and site-directed mutagenesis, the mutation does not affect the binding affinity of the SepSecS-tRNA complex. The mutant forms a stable complex with GroEL. The side chain of Tyr334 is in helix alpha13 near the active-site pocket. Its hydroxyl group forms a hydrogen bond with the backbone carbonyl of Asn285, and this interaction may help stabilize a loop that carries Lys284 and the covalently attached PLP cofactor. In the Y334C crystal, the side chain of Cys334 coordinates two water molecules, which interact with the backbone carbonyl of Asn285 in the same fashion as the Tyr side chain in the wild-type enzyme. Tetramers of Y334C adopt the same structure as wild-type SepSecS. The pathogenic mutation Tyr334Cys does not alter the three-dimensional structure of the SepSecS tetramer

Y429*

naturally occurring nonsense mutation and site-directed mutagenesis. Y429* expresses at low levels and as insoluble protein regardless of the incubation temperature, induction point, or the growth media used. Tyr429 is located before strand beta14. Premature abortion of protein synthesis yields a truncated enzyme devoid of strand beta14, loop beta14-alpha15, and the C-terminal helix alpha15. Loop beta14-alpha15 establishes a side of the catalytic groove, and helix alpha15 provides residues that bind the 5'-end of tRNASec. The Y429* variant is not be capable of promoting selenocysteine synthesis

H166A

the mutant is partially active in forming Sec-tRNASec in vivo. In vitro, the mutant is partially active in forming Cys-tRNASec

H166F

mutant is inactive in vivo

H166Q

mutant is inactive in vivo

R307A

the mutant is significantly less active in L-selenocysteinyl-tRNASec formation in vivo and Cys-tRNASec formation in vitro

R72A

the mutant enzyme is significantly less active in L-selenocysteinyl-tRNASec formation in vivo and Cys-tRNASec formation in vitro. The mutant enzyme is unable to form L-selenocysteinyl-tRNASec in vitro

additional information

recombinant enzyme

3 entries

recombinant His6-tagged enzyme mutants from Escherichia coli strain BL21(DE3) and SoluBL21(DE3) by nickel affinity chromatography

recombinent enzyme

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3 entries

expressed in Escherichia coli SelA-deficient strain JS1

Leishmania donovani

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738683

gene SEPSECS, recombinant expression of His6-tagged enzyme mutants in Escherichia coli strain BL21(DE3) and SoluBL21(DE3) (a bacterial strain engineered to increase solubility of recombinant proteins)

phylogenetic analysis and tree