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spinqualitylabelsall models Crystal structure of firefly luciferase Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Biological context Firefly, also named Photinus pyralis, is a bioluminescent insect. They are able to produce light by an energetic process, in order to attract its mate. The enzyme responsible of this light producing is luciferase, also known as luciferin-4-monooxygenase (EC: 1.13.12.7). It acts also as a ligase. This enzyme has various applications in the biotechnology field. In fact, it is used in chemical biology and drug trials. As there is no light production by mammals, luciferase is really a great tool for researchers. But this light emission depends on the environmental conditions. For example, they help to detect protein-protein interactions, to track cells in vivo in order to analysis the development of disease at the molecular level in real-time , to monitor the transcriptional and post-transcriptional regulation of specific gens, to control apoptosis, to label cancer cells, to detect environmental contaminations … Global Structure Luciferase is a 62kDa protein. It contains 550 amino acids. The enzyme can be divide into two domains. On the one hand, the major portion, corresponding to the N-terminal. On the other hand, the small portion, corresponding to the C-terminal domain.Those two domains are separated by a large cleft. The C terminal domain The amino acids of C terminal sequence is composed of contiguous residues and form a type of lid upon the N terminal domain. It contains two β sheets: the first β sheet is composed of two short antiparallel strands and the second β sheet is composed of 3 antiparallel strands, mixed with three α helices. Those helices are put toward the outside.It is α + β structure. The N terminal domain The amino acids of C terminal sequence is composed of non contiguous residues. It falls into three subdomains (the subdomain A,the subdomain B, and the subdomain C).It has an antiparallel β barrel, two β sheets, which are framed by α helices. Each of the two β sheets subdomains (A and B)are composed of 8 β strands and 6 helices. The β sheet A has 5 parallel and 3 antiparallel β strands. The β sheet B has 6 parallel and 2 antiparallel β strands. Those β sheets create a groove, closed on one end by the β barrel. Structure related to functions The most known reaction of luciferase is the light emission where luciferase uses luciferin, ATP and O2 as substrates. The color of emitted light varies according to the pH, which can be explained by the luciferase structure. But there are some other reactions which can uses fatty acids and coenzyme A. So the active site of the luciferase can theorically bind all these compounds. Interactions with ligands The active site is not strictly highlighted according to the actual state of studies but some residues and motifs strongly modified have been determined and this conformation enables to find the active site. Many of these conserved residue are located on the core of the β-barrel and on the small C-terminal domain and in the surface of the N-terminal domain, which forms a depression. However, this depression is too large to enable interactions between residues and substrates, so it is thought that a conformational change occurs and sandwiches the substrates, forming the active site. This conformational change provide a suitable environment for light production because of water molecules will be excluded from the active site, favouring intramolecular reactions. Residues also follow a cleft caused by of the Beta-sheet B against the Beta-barrel. Hydrogen bonding between Luciferase and substrates luciferin (green), ATP (violet) and Mg2+, Interaction with ATP We find a signal motif in luciferase which is [STG]-[STG]-G-[ST]-[ST]-[TSE]-[GS]-x-[PALIVM]-K where some residue like lysine are always conserved. This pattern enables ATP binding thanks to hydrogen bonds between residues and phosphates of ATP. There is another pattern : [YFW]-[GASW]-x-[TSA]-E which takes a particular conformation because of hydrogen bonds between residues and maintain the adenosin ring of ATP., Interaction with luciferin Luciferase holds the luciferin with the specific residues arginin 218, phenylalanin 247, serin 347 and adenin 348, still with hydrogen bounds. This bindings makes the carboxylate oxygen of luciferin points toward the α phosphate of ATP, so the oxygen is well-positionned to attack the α phosphate. This promotes the luciferin-AMP formation., Interaction with fatty acids Fatty acids are highly similar to luciferin. Therefore, luciferase can use the luciferin binding site to bind fatty acids. That is why they can be used as substrates by luciferase and then, very high similar reaction as for luciferin occurs. Comparison of the chemical structures of (A) firefly D-LH2 and (B) arachidonic acid, Color modulation When the pH is low, the color of light changes. This is probably due to the hydrogen bonds network between substrates, residues of the cleft and water. Indeed, this network triggers to an external electrostatic potential which stabilizes a charge created during the reaction of light emission. The lower pH leads to a weakening of the network, so the energy which can be emitted decreases and the wavelength of photon increases and becomes more red. Evolution The number of proteins related to luciferase is growing exponentially. They are a lot of enzymes very different involved in a lot of mechanisms such as biosynthesis of siderophores, of antibiotics, fatty acid:coenzyme A ligase, and so on... All the coenzyme A ligase show a very high level of similarity : indeed, each of these enzyme related to luciferase catalyze the adenylation of a carboxylic acid substrate using ATP-Mg2+ and then, the ligation of the activated carboxylic acid with an acceptor. That is why luciferase is more and more considered as coming from a common ancestor involved in this kind of reaction and the light production would only be a side effect of the reaction.