Literature summary for 2.9.1.2 extracted from

Puppala, A.K.; French, R.L.; Matthies, D.; Baxa, U.; Subramaniam, S.; Simonovic, M.
Structural basis for early-onset neurological disorders caused by mutations in human selenocysteine synthase (2016), Sci. Rep., 6, 32563 .

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Cloned(Commentary)

Cloned (Comment)	Organism
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)	Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
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	Homo sapiens
additional information	mapping pathogenic mutations onto the tetrameric human SepSecS-tRNASec complex, overview	Homo sapiens
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	Homo sapiens
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	Homo sapiens
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	Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
O-phospho-L-seryl-tRNASec + selenophosphate + H2O	Homo sapiens	-	L-selenocysteinyl-tRNASec + 2 phosphate	-	?

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	Q9HD40	-	-

Purification (Commentary)

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

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
additional information	SepSecS binds unacylated tRNASec equally well as Sep-tRNASec 1	Homo sapiens	?	-	-
O-phospho-L-seryl-tRNASec + selenophosphate + H2O	-	Homo sapiens	L-selenocysteinyl-tRNASec + 2 phosphate	-	?

Subunits

Subunits	Comment	Organism
homotetramer	4 * 60000, recombinant enzyme, SDS-PAGE	Homo sapiens

Synonyms

Synonyms	Comment	Organism
O-phosphoseryl-tRNASec:selenocysteinyl-tRNASec synthase	-	Homo sapiens
SepSecS	-	Homo sapiens

General Information

General Information	Comment	Organism
malfunction	four distinct mutations (A239T, Y334C, T325S and nonsense Y429*) in human gene SEPSECS cause congenital cerebellar atrophy termed pontocerebellar hypoplasia type 2D (PCH2D). Pontocerebellar hypoplasia (PCH) is a group of autosomal recessive disorders affecting different cerebral structures, particularly the brainstem and cerebellum. Most PCH types result from mutations in genes important for tRNA splicing and aminoacylation and RNA transport. The PCH2D patients similarly suffer from progressive cerebellar and cerebral atrophy, neonatal irritability, and debilitating spasticity. Neuropathological analysis reveals severe atrophy of the brainstem and cerebellar cortex with loss of both white and gray matter. This subset of patients also exhibits a slight reduction in selenoprotein levels, suggesting that SepSecS catalysis is impaired. Pathogenic variants are less soluble than wild-type SepSecS. Mutations Thr325Ser and Tyr334Cys do not affect the binding affinity of the SepSecS-tRNA complex	Homo sapiens
physiological function	the enzyme is responsible for the formation of only 25 human proteins, but the human selenoproteome is pivotal for the maintenance of the cellular redox potential (e.g. thioredoxin reductases), regulation of the overall metabolic rate (e.g. iodothyronine deiodinases), removal of reactive oxygen species and prevention of oxidative damage (e.g. glutathione peroxidases, and methionine sulfoxide reductases), and selenium homeostasis (e.g. selenoprotein P)	Homo sapiens

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Release 2023.1 (January 2023)