Abstracts

Ca²⁺-regulated photoproteins (CaPhs), to which the prosthetic group coelenterazine binds in the protein’s internal cavity, undergo conformational changes uppon Ca²⁺ addition, causing emission of light. Recently, we reported a novel class of CaPhs - velamins - isolated from the bioluminescent ctenophore Velamen parallelum. In contrast to all other CaPhs, that produce light within the blue range (λ_max 465-495 nm), velamins are capable of emitting green light (λ_max 500-508 nm).CaPhs from hydromedusae (aequorin, mitrocomin, clytin, and obelin) and ctenophores (mnemiopsin, berovin, bolinopsin, and bathocyrovin) have been extensively investigated lately. Despite sharing some common structural features, including calcium-binding loops and substrate, ctenophore and hydromedusae photoproteins have low primary sequence identity and differ on certain intrinsic properties. For example, photoinactivation is present in ctenophores while hydromedusae photoproteins have a higher sensitivity towards calcium. Phylogenetic analysis of V. parallelum CaPhs revealed three functionally diverse clusters of photoproteins: whilst α-velamin isoforms exhibited the highest light emission activity, β- and γ-velamins were found to be more thermostable at higher temperatures. By comparing velamin sequences to other CaPhs, we found a strictly conserved ESYRYLRS motif located at the N-terminus of all ctenophore CaPhs and which is absent in hydromedusan counterparts. Here, we demonstrate that the deletion of the N-terminus alfa-helix motif in α-velamin isoform ablates its function, revealing its relevance for the bioluminescence of ctenophores. Further in silico modelling studies may provide new insights on the role of these residues in CaPhs, specially regarding the binding of coelenterazine and calcium ions. These findings reveal a robust phylogenetic marker to support the two different CaPhs classes from hydromedusae and ctenophores, as well as they contribute to our understanding of the ecology and evolution of bioluminescence in marine organisms.