Cysteine-390 is the binding site of luminous substance with symplectin, a photoprotein from Okinawan squid, Symplectoteuthis oualaniensis.
Symplectin is a photoprotein from a luminous squid, Symplectoteuthis oualaniensis. It has a luminous substrate, dehydrocoelenterazine (DCZ), linked through a thioether bond with a cysteine residue. We have proven the binding site of luminous substrate in symplectin by using an artificial analogue of DCZ, ortho-fluoro-DCZ (F-DCZ). F-DCZ-symplectin emitting strong blue light was reconstituted from apo-symplectin and F-DCZ. Proteolytic digestion of the reconstituted F-DCZ-symplectin afforded peptides including C(390)GLK-F-DCZ (amide), which was detected with a house assembled nano-LC-ESI-Q-TOF-MS. The chromo-peptide derived from the F-DCZ-symplectin after luminescence showed the lower molecular mass than that before the luminescence by 12 mass units, corresponding to the loss of one carbon atom upon emitting light. Thus, we have concluded that F-DCZ analogue binds to Cys390 in symplectin so as to emit light.
Isobe
Minoru
M
Institute for Advanced Research, Nagoya University, Aichi, Japan. isobem@ff.iij4u.or.jp
Kuse
Masaki
M
Tani
Naoki
N
Fujii
Tatsuya
T
Matsuda
Tsukasa
T
D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain; D003545:Cysteine / Q000378:metabolism*; D049832:Decapodiformes / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D049449:Luminescence; D049408:Luminescent Agents / Q000378:metabolism*; D008164:Luminescent Proteins / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data
D049408:Luminescent Agents; D008164:Luminescent Proteins; D003545:Cysteine