Molecular evolution of dinoflagellate luciferases, enzymes with three catalytic domains in a single polypeptide

Abstract

Enzymes with multiple catalytic sites are rare, and their evolutionary significance remains to be established. This study of luciferases from seven dinoflagellate species examines the previously undescribed evolution of such proteins. All these enzymes have the same unique structure: three homologous domains, each with catalytic activity, preceded by an N-terminal region of unknown function. Both pairwise comparison and phylogenetic inference indicate that the similarity of the corresponding individual domains between species is greater than that between the three different domains of each polypeptide. Trees constructed from each of the three individual domains are congruent with the tree of the full-length coding sequence. Luciferase and ribosomal DNA trees both indicate that the Lingulodinium polyedrum luciferase diverged early from the other six. In all species, the amino acid sequence in the central regions of the three domains is strongly conserved, suggesting it as the catalytic site. Synonymous substitution rates also are greatly reduced in the central regions of two species but not in the other five. This lineage-specific difference in synonymous substitution rates in the central region of the domains correlates inversely with the content of GC 3, which can be accounted for by the biased usage toward C-ending codons at the degenerate sites. RNA modeling of the central region of the L. polyedrum luciferase domain suggests a function of the constrained synonymous substitutions in the circadian-controlled protein synthesis.codon bias ͉ domain duplication ͉ synonymous substitutions T he luciferase (LCF) of the bioluminescent dinoflagellate Lingulodinium polyedrum (Lp; formerly Gonyaulax polyedra) has three homologous tandemly arranged 377-aa-long domains, each of which is catalytically active (1). The LCF of another dinoflagellate, Pyrocystis lunula (Pl), has a similar structure (2). In both, we found the amino acids in the central region of each of the three domains to be highly conserved intramolecularly at Ϸ95% identity, compared with Ϸ65% in the flanking regions. Remarkably, and only in Lp, there were fewer synonymous substitutions in the central region.These results are intriguing from both evolutionary and mechanistic viewpoints. To grasp their significance, we investigated the LCFs of five other bioluminescent dinoflagellates, all of which are photosynthetic: Alexandrium affine (Aa), Alexandrium tamarense (At), Pyrocystis fusiformis (Pf ), Pyrocystis noctiluca (Pn), and Protoceratium reticulatum (Pr). All were shown to have the same basic configuration, i.e., tandem triplication of catalytic domains following an Ϸ110-aa-long N-terminal sequence. Fortuitously, it turned out that Lp and Pl LCFs represent extremes of a spectrum of LCF structures with regard to synonymous substitution rates in the central region of each domain.Bioluminescence in dinoflagellates has been well characterized only in Lp, where the three components involved in light emission are packaged together in unique cellular organelles,...

Two different domains of the luciferase gene in the heterotrophic dinoflagellate Noctiluca scintillans occur as two separate genes in photosynthetic species

“…Previous studies revealed that the amount of bias in codon usage varies in the central regions of the domains of the luciferases of the seven photosynthetic dinoflagellates (9,10). The effective numbers of codons is a measure of codon biases (higher value, lower bias).…”

“…In six other photosynthetic dinoflagellates, the structures of the luciferases were found to be essentially the same as in Lp, and all were found to occur in multiple and tandem copies, albeit with very different intergenic sequences (9,10). In some of these species, a luciferin-binding protein has not yet been found, but in vitro, all react to emit light with the luciferin of Pyrocystis lunula (Pl), which was established to be an open tetrapyrrole (11) and will be called dinoflagellate luciferin.…”

“…The selection of specific parameters for the tree construction and other bioinformatic analysis were done as described in ref. 9.…”

Dramatic blooms ofPrymnesiumsp. (Prymnesiophyceae) andAlexandrium margalefii(Dinophyceae) in the Salton Sea, California

“…Dinoflagellates are the only photosynthetic organisms capable of exhibiting bioluminescence (Sweeney 1987), and a number of genera of marine dinoflagellates have been shown to produce light, including Alexandrium Halim. Species within this genus known to be bioluminescent are A. fundyense (Balech) Balech and A. tamarense (Lebour) Balech (Anderson et al 1994), A. affine (Liu et al 2004) and A. catenella (Sullivan and Swift 2003), however, not all clones in a species produce bioluminescence (Anderson et al 1994).…”

Profile of J. Woodland Hastings

“…So more active sites without more molecules,'' he explains. Postdoctoral fellow Liyun Liu examined luciferases from seven different dinoflagellate species and found them all to be like Gonyaulax: an N-terminal region of unknown function followed by three homologous luciferase domains (23).…”