User:Matt Whelihan: Difference between revisions

==Luciferases==

Bioluminescence is the process by which living organisms convert chemical energy into photons of light and it is widely distributed throughout the animals, plants and fungi <ref>PMID:6358519</ref>. Species use bioluminescence as a survival tool in mating, defense and hunting. The one thing that all bioluminescent species have in common is that they all catalyze the reaction with an enzyme generically called a luciferase.  All luciferases oxidize a substrate, which then decays back to the ground state while emitting a photon of light. This process is incredibly efficient with almost one photon of light produced per oxidation. While all lucifeases oxidize their substrates, the cofactors involved and reaction pathways used, vary widely <ref>PMID:775940</ref><ref>PMID:14444706</ref>.  

Luciferases have become an invaluable tool in microbiology and biochemistry as a means of reporting gene expression and in-vivo/vitro chemical conditions. Since it was first cloned in 1985 by DeLuca et al., the gene coding for luciferases has been used in reporter assays to measure gene transcription and cellular morphology. Various cell lines have also been engineered to express luciferases as a measure of the oxidative state in various organs and types of diseases. Perhaps its most useful function is in the efficient detection of intracellular ATP. Luciferases are also used to detect protein-anesthetic interactions as they are susceptible to a wide range of general anesthetics.   

Both the Japanese and North American luciferase variants consist of a single-chain 62 kDa monooxygenases with two distinct domains  that are <scene name='User:Matt_Whelihan/Secondary_structure/1'>comprised</scene> of a mix of <font color='magenta'>alpha</font> and <font color='gold'>beta</font> secondary structures. The <scene name='User:Matt_Whelihan/N-terminal_domain/1'>N-terminal domain</scene> (residues 4-436) is comprised of of an antiparalell β-barrel and two β-sheets, flanked by α-helices which forms a typical <scene name='User:Matt_Whelihan/Rossmann/1'>Rossmann fold</scene> [http://images.google.com/imgres?imgurl=http://chem.csusb.edu/~dpedersn/C436/openbetarossman.jpg&imgrefurl=http://chem.csusb.edu/~dpedersn/C436/prot_struct_and_class.html&usg=__xOF4ifmAyJ5qm3UPaNKd9whKlu4=&h=539&w=719&sz=35&hl=en&start=5&um=1&tbnid=IMkWKmy0xzWcYM:&tbnh=105&tbnw=140&prev=/images%3Fq%3DRossman%2BFold%26hl%3Den%26safe%3Doff%26sa%3DN%26um%3D1] The  

<scene name='User:Matt_Whelihan/C-terminal_domain/2'>C-terminal domain</scene> (residues 440-544) is comprises of a separate α-β hinge<ref>PMID:8805533</ref>. Firefly luciferases share significant sequence and mechanistic homology with peptide synthetases and acylCoA ligases. These enzymes belong to a superfamily of adenylate-forming enzymes that catalyze activation reactions between ATP and a carboxyl group of their substrates. This group of proteins shares an identifying motif <scene name='User:Matt_Whelihan/198-207/1'>(198SerSerGlySerThrGlyLeuProLysGly207)</scene> and has been termed the “acyl-adenylate/thioester-forming” enzyme family. Despite high sequence homology, there are only seven residues that are conserved across this superfamily (Gly200, Lys206, Glu344, Asp422, Arg437, Gly446, and Glu455). These residues are believed to be integral to the binding of ATP and the formation of an adenylate compound <ref>PMID:1351742</ref><ref>PMID:1447981</ref>. These residues however are located across the N and C-terminal domains, which in the structure, are too far apart to produce catalysis. This suggested suggest that the crystallized form was in the resting state of the enzyme and it was hypothesized that the C-terminal domain would close in on the active site cleft upon substrate binding. This closing of the active site combined with various hydrophobic residues seen packed around the active site suggested that catalysis may occur in the absence of water.

== Catalysis ==