SEPHS2

SEPHS2 is a selenophosphate synthetase that catalyzes the ATP-dependent conversion of selenide to selenophosphate, the essential selenium donor for selenocysteine biosynthesis. The enzyme employs an active-site selenocysteine residue (or a functional cysteine surrogate) that is absolutely required for catalysis, as demonstrated by genetic complementation of E. coli selD mutants and in vitro kinetic analysis (PMID:9012797, PMID:10515607); SEPHS2 preferentially assimilates inorganic selenite as its selenium source, distinguishing it functionally from SEPHS1 (PMID:15534230). Within mammalian cells, SEPHS2 physically associates with SEPSECS and SEPHS1 in oligomeric complexes that constitute the selenocysteine incorporation machinery (PMID:28414460). Independent of selenoprotein biosynthesis, SEPHS2 loss suppresses oxidative phosphorylation and elevates intracellular NAD⁺, activating the deacetylase SIRT2 to stabilize the gluconeogenic enzyme PCK1 and redirect glucose flux toward the pentose phosphate pathway (PMID:42035418).

1. 1997 High

   Establishing that mammalian SPS2 (SEPHS2) possesses intrinsic selenophosphate synthetase activity and that the active-site selenocysteine (or its Cys surrogate) is catalytically competent answered the fundamental question of whether mammals encode their own selenophosphate-generating enzyme.

   Evidence Baculovirus-expressed purified Cys-mutant enzyme assayed for selenide-dependent AMP formation from ATP

   PMID:9012797

   Open questions at the time

   • Kinetic parameters for the native Sec-containing enzyme were not determined
   • In vivo role in mammalian selenium metabolism not yet demonstrated
2. 1999 High

   Demonstrating that SPS2 functionally complements bacterial selD deficiency and that only Cys/Sec at the active site supports activity resolved which residue is catalytically essential and provided kinetic constants (Km(ATP) = 0.75 mM).

   Evidence Site-directed mutagenesis (Cys→Ala/Ser/Thr), complementation of E. coli selD mutant, purified enzyme kinetics

   PMID:10515607

   Open questions at the time

   • No structural model explaining how the active-site chalcogen participates in catalysis
   • Role of SEPHS1 versus SEPHS2 not yet distinguished
3. 2004 High

   Distinguishing SEPHS2 from SEPHS1 by substrate specificity—SEPHS2 preferentially uses inorganic selenite while SEPHS1 relies on a selenocysteine lyase salvage route—resolved a longstanding question about functional redundancy between the two paralogs.

   Evidence Complementation of E. coli selD mutant with selenite vs. L-selenocysteine, formate dehydrogenase H activity readout

   PMID:15534230

   Open questions at the time

   • Mechanism by which selenite is reduced to selenide upstream of SEPHS2 not defined
   • Whether SEPHS2 contributes to selenium salvage under physiological conditions unclear
4. 2017 Medium

   Identifying physical interactions between SEPHS2, SEPSECS, and SEPHS1 established that selenophosphate synthesis and selenocysteine biosynthesis occur within a multi-protein complex rather than via freely diffusing intermediates.

   Evidence BRET and co-immunoprecipitation in mammalian cells

   PMID:28414460

   Open questions at the time

   • Stoichiometry and architecture of the complex not determined
   • Functional consequence of complex disruption not tested
   • Single study; independent replication pending
5. 2025 Medium

   Placing SEPHS2 translation downstream of METTL5-mediated 18S rRNA m6A modification revealed a translational control layer for selenoprotein biosynthesis and explained how METTL5 loss causes selenoprotein deficiency and ROS-mediated apoptosis in myeloma cells.

   Evidence METTL5 KO/KD in myeloma cells and orthotopic xenografts, selenoprotein synthesis and ROS assays

   PMID:40750759

   Open questions at the time

   • Whether SEPHS2 mRNA has specific structural features conferring METTL5 sensitivity is unknown
   • Generalizability beyond multiple myeloma not tested
6. 2026 High

   Discovering that SEPHS2 loss suppresses OXPHOS and redirects metabolism through NAD⁺/SIRT2-dependent PCK1 stabilization toward the pentose phosphate pathway established a non-canonical, selenoprotein-independent metabolic function for SEPHS2.

   Evidence OXPHOS genetic screen, SEPHS2 KO, metabolic flux analysis, NAD⁺ measurement, SIRT2 activity and PCK1 acetylation assays, in vivo tumor model

   PMID:42035418

   Open questions at the time

   • How SEPHS2 mechanistically controls NAD⁺ levels remains undefined
   • Whether the metabolic phenotype reflects a moonlighting enzymatic activity or loss of a metabolite is unresolved
   • Relevance in non-cancer tissues not examined

• The mechanism by which SEPHS2 regulates intracellular NAD⁺ independently of selenoprotein biosynthesis, and whether SEPHS2 has additional non-canonical substrates or enzymatic activities, remain open questions.
• No structural model of human SEPHS2 explaining dual functionality
• Upstream selenium delivery pathway to SEPHS2 in mammalian cells not fully reconstituted
• The relationship between selenophosphate synthesis and NAD⁺ metabolism is mechanistically unresolved