The bioluminescence of many marine coelenterates involves the interaction of two proteins, a Ca²⁺-regulated photoprotein in the jellyfish case, aequorin, and its cognate green-fluorescent protein, Aequorea GFP. Addition of Ca²⁺to the purified aequorin produces a blue bioluminescence. It was early recognized that, in the jellyfish itself, the in vivo bioluminescence was a green color and after further study, the origin of this green emission was identified as the GFP. How is this bioluminescence spectrum shifted?

Förster resonance energy transfer within a protein-protein complex has previously been invoked to explain emission spectral modulation.However, the well-known Förster theory requires concentrations of the donor-acceptor partners in the millimolar range, whereas in some bioluminescence systems, the GFP effect on the in vitro bioluminescence is observed at micromolar concentrations.

In a recent study published in the Journal of Biological Chemistry, 285, 40891–40900, Dr. LIU Zhijie and his colleagues at the Institute of Biophysics, Chinese Academy of Sciences presented a spatial structure of a complex of the Ca²⁺-regulated photoprotein clytin with its green-fluorescent protein (cgGFP) from the jellyfish Clytia gregaria, and showed that it accounted for the bioluminescence properties of this system in vitro. Based on the structures, the interaction surfaces in the complex were identified by NMR titration experiments. Perturbation of chemical shifts of the separate proteins, which could be mapped to particular residues, was affected by complexation. The association was weak but from knowledge of the interaction surface. The overall structure of the complex was modeled using computational docking approach. To verify the clytin-cgGFP structure they performed site-directed mutagenesis of clytin residues located at the interaction site which could reduce the degree of protein-protein association concomitant with a loss of effectiveness of cgGFP in color-shifting the bioluminescence. The clytin-cgGFP structure may correspond to the transient complex postulated to account for the energy transfer effect of GFP in bioluminescence systems of Aequorea and Renilla, aiding an understanding of how GFP functions in vivo.

FIGURE . Stereoview representation of the spatial structure of the clytin-cgGFP complex derived from x-ray structures of clytin and cgGFP, NMR-mapping of the interaction surfaces and computational docking in HADDOCK. 45 Å is the distance between the two chromophores. Structural elements of clytin and cgGFP comprising the interaction surface are labeled. (From Dr. LIU Zhijie )