Identified a 60 kDa photoprotein from S. oualaniensis photogenic organ, extracted with 0.6 M KCl as luminescence-active forms.

Protein exists mainly as oligomers (~200 kDa) with trace monomer; partial tryptic digestion yields a 40 kDa luminescent fragment and 16 kDa N-terminal fragment.

Amino acid sequencing revealed no similarity to known photoproteins but significant similarity to the carbon-nitrogen hydrolase domain of mammalian biotinidase and vanin (pantetheinase).

Immunoblot analysis showed specific expression of the 60 kDa protein in the photogenic organ.

Proved that Cys-390 is the binding site for dehydrocoelenterazine (dhCtz) via a thioether bond using fluorinated dhCtz analogs and nano-LC-ESI-Q-TOF-MS.

Light emission requires monovalent cations (K+ or Na+) and O2 at optimum pH 7.8; the reaction releases CO2 (loss of 12 mass units).

Symplectin emits blue light at 470 nm.

The chromophore is covalently bound and cannot be extracted by normal solvent procedures.

Cephalopods have four well-supported paralog groups of the biotinidase/pantetheinase protein family; symplectin forms one group.

The Cys-390 homolog (dhCtz binding site) is conserved across essentially all members of the protein family.

Active site residues for pantetheinase catalysis (E60, K163, C196 in symplectin) are also conserved, suggesting symplectin may retain hydrolase activity and have dual function.

Symplectin homologs found in non-luminous species (Sepia pharaonis, Loligo vulgaris, Doryteuthis pealei), indicating tree position alone does not predict bioluminescence.

The protein structure is modeled as two domains: the nitrilase domain (residues 1-290) and the base domain (291-465), with bioluminescence activity in the base domain near Cys-390.

Demonstrated that stereochemistry at the Cys-390 binding site is dynamic and plays a critical role in bioluminescence efficiency.

Two fluorinated dhCtz analogs (2,4-diF-DCL and 2,6-diF-DCL) showed dramatically different luminescence activities (200% and 20% of natural dhCtz, respectively).

Identified coelenteramide-390-CGLK-peptide as a product of the luminescence reaction.

Demonstrated dynamic exchange of chromophores at the binding sites and movement to the active site Cys-390 for luminescence.

Showed that symplectin can reconstitute with various dhCtz analogs at pH 6.0 and subsequently luminesce at pH 7.8.

Identified a membrane-bound ~60 kDa photoprotein from D. gigas photophores that uses dehydrocoelenterazine and emits 470 nm blue light.

LC/MS analysis matched the photoprotein to symplectin and vanin-2 gene products from D. gigas transcriptome with >80% coverage.

Reviewed the structural analysis of chromophores in photoproteins of the flying squid and mollusk, formed by connecting apo-protein with dehydrocoelenterazine.

The photogenic organ produces an intense blue flash followed by rapid decay, with kinetic analysis suggesting two light-emitting components

Monovalent cations trigger luminescence: K+ is most effective, followed by Rb+, Na+, Cs+, NH4+, and Li+; divalent cations (Ca2+, Mg2+, Sr2+) do not trigger light emission

The essential light-emitting components are membrane-bound within granules in the photogenic organ

Optimal activation requires 0.6 M KCl or NaCl at pH 7.8; a free sulfhydryl group is essential for activity

S. oualaniensis is among the studied species for cephalopod bioluminescence, using an intrinsic enzyme (symplectin) rather than symbiotic bacteria

Squids have evolved multiple strategies for bioluminescence, including symbiotic bacteria in some species and intrinsic enzymes like symplectin in others