View source for Luciferase

{{Infobox protein family
| Symbol = Bac_luciferase
| Name = Bacterial Luciferase monooxygenase family
| width =
| Pfam = PF00296
| InterPro =IPR016048
| SMART =
| PROSITE =PDOC00397
| SCOP =1nfp
| TCDB =
| OPM family =
| OPM protein =
| PDB = {{PDB2|1brl}}, {{PDB2|1bsl}}, {{PDB2|1ezw}}, {{PDB2|1fvp}}, {{PDB2|1luc}}, {{PDB2|1m41}}, {{PDB2|1nfp}}, {{PDB2|1nqk}}, {{PDB2|1rhc}}, {{PDB2|1xkj}}
}}
{{Infobox protein family
| Symbol = Luciferase_cat
| Name = Dinoflagellate Luciferase catalytic domain 
| image = PDB 1vpr EBI.jpg
| width =
| caption = crystal structure of a luciferase domain from the dinoflagellate ''Lingulodinium polyedrum''
| Pfam = PF10285
| Pfam_clan =
| InterPro = IPR018804
| SMART =
| PROSITE =
| MEROPS =
| SCOP =
| TCDB =
| OPM family =
| OPM protein =
| CAZy =
| CDD =
}}
{{Infobox protein family
| Symbol = Luciferase_N
| Name = Dinoflagellate Luciferase/LBP N-terminal domain
| image =
| width =
| caption =
| Pfam = PF05295
| Pfam_clan =
| InterPro = IPR007959
| SMART =
| PROSITE =
| MEROPS =
| SCOP =
| TCDB =
| OPM family =
| OPM protein =
| CAZy =
| CDD =
}}
{{Infobox protein family
| Symbol = Luciferase_3H
| Name = Dinoflagellate Luciferase helical bundle domain
| image = NONE
| width =
| caption = 
| Pfam = PF10284
| Pfam_clan =
| InterPro = IPR018475
| SMART =
| PROSITE =
| MEROPS =
| SCOP =
| TCDB =
| OPM family =
| OPM protein =
| CAZy =
| CDD =
}}
{{Infobox nonhuman protein
| Name = Firefly luciferase
| caption = Structure of ''Photinus pyralis'' [[firefly]] luciferase.
| image = Firefly_Luciferase_Crystal_Structure.rsh.png
| width = 200
| Organism = ''[[Photinus pyralis]]''
| TaxID = 7054
| HGNCid =
| Symbol = Firefly luciferase
| AltSymbols =
| EntrezGene =
| OMIM =
| RefSeq =
| UniProt = P08659
| PDB = 1LCI
| PDB_supplemental = [http://www.ebi.ac.uk/pdbe/searchResults.html?display=both&term=P08659 More structures]
| ECnumber = 1.13.12.7
| Chromosome =
| Arm =
| Band =
| LocusSupplementaryData =
}}
'''Luciferase''' is a generic term for the class of oxidative [[enzyme]]s that produce [[bioluminescence]], and is usually distinguished from a [[photoprotein]]. The name was first used by [[Raphaël Dubois]] who invented the words ''[[luciferin]]'' and ''luciferase'', for the substrate and [[-ase|enzyme]], respectively.<ref>{{Cite web | vauthors = Lee J | title = A History of Bioluminescence | date = 28 February 2014 | url = http://photobiology.info/HistBiolum/HistBiolum.html | url-status = live | archive-url = https://web.archive.org/web/20150329013414/http://photobiology.info/HistBiolum/HistBiolum.html | archive-date = 29 March 2015 | website = photobiology.info }}</ref> Both words are derived from the Latin word ''lucifer'', meaning "lightbearer", which in turn is derived from the Latin words for "light" (''lux)'' and "to bring or carry" (''ferre)''.<ref>{{Cite journal | vauthors = Lee J | title = Bioluminescence: the First 3000 Years (Review) | journal = Journal of Siberian Federal University. Biology | volume = 1 | issue = 3 | pages = 194–205 | date = September 2008 | doi = 10.17516/1997-1389-0264 | url = https://www.researchgate.net/publication/229052741 | doi-access = free }}</ref>Luciferases are widely used in [[biotechnology]], for [[bioluminescence imaging]]<ref>{{cite journal | vauthors = Brennan CK, Ornelas MY, Yao ZW, Prescher JA | title = Multicomponent Bioluminescence Imaging with Naphthylamino Luciferins | journal = ChemBioChem | volume = 22 | issue = 16 | pages = 2650–2654 | date = August 2021 | pmid = 34139065 | pmc = 8496354 | doi = 10.1002/cbic.202100202 }}</ref> microscopy and as [[reporter gene]]s, for many of the same applications as [[fluorescent protein]]s. However, unlike fluorescent proteins, luciferases do not require an external [[Fluorescence|light source]], but do require addition of [[luciferin]], the consumable substrate.

== Examples ==
A variety of organisms regulate their light production using different luciferases in a variety of light-emitting reactions. The majority of studied luciferases have been found in animals, including [[fireflies]],<ref>{{cite web | title = X-Shining Thermostable Luciferase | url = https://www.biosynth.com/scientific-resources/product-guides/x-shining-thermostable-luciferase }}</ref> and many marine animals such as [[copepods]], [[scyphomedusa|jellyfish]], and the [[Renilla reniformis|sea pansy]]. However, luciferases have been studied in luminous fungi, like the [[Omphalotus olearius|Jack-O-Lantern mushroom]], as well as examples in other kingdoms including [[bioluminescent bacteria]], and [[dinoflagellates]].

=== Firefly and click beetle ===

The [[Firefly luciferase|luciferases of fireflies]] – of which there are over 2000 [[species]] – and of the other [[Elateroidea]] (click beetles and relatives in general) are diverse enough to be useful in [[molecular phylogeny]].<ref>{{cite journal | vauthors = Gould SJ, Subramani S | title = Firefly luciferase as a tool in molecular and cell biology | journal = Analytical Biochemistry | volume = 175 | issue = 1 | pages = 5–13 | date = November 1988 | pmid = 3072883 | doi = 10.1016/0003-2697(88)90353-3 }}</ref>  In fireflies, the oxygen required is supplied through a tube in the abdomen called the [[abdominal]] [[Invertebrate trachea|trachea]].  One well-studied luciferase is that of the [[Photinini]] firefly ''[[Photinus pyralis]]'', which has an optimum pH of 7.8.<ref>{{cite journal | vauthors = Steghens JP, Min KL, Bernengo JC | title = Firefly luciferase has two nucleotide binding sites: effect of nucleoside monophosphate and CoA on the light-emission spectra | journal = The Biochemical Journal | volume = 336 | issue = Pt 1 | pages = 109–113 | date = November 1998 | pmid = 9806891 | pmc = 1219848 | doi = 10.1042/bj3360109 }}</ref>

[[Click beetle]]<ref>{{Cite journal | vauthors = Klapach A, Lorenz WW, Tsai LK, Laughlin LA, Gorman JA, Soll DR, Srikantha T | title = The sea pansy Renilla reniformis luciferase serves as a sensitive bioluminescent reporter for differential gene expression in Candida albicans | journal = Journal of Bacteriology | volume = 178 | issue = 1 | pages = 121–129 | date = January 1996 | pmid = 8550405 | pmc = 177628 | doi = 10.1128/jb.178.1.121-129.1996 | language = en | issn = 0021-9193 }}</ref>

=== Sea pansy ===

Also well studied is the sea pansy, ''[[Renilla reniformis]]''. In this organism, the luciferase ([[Renilla-luciferin 2-monooxygenase]]) is closely associated with a luciferin-binding protein as well as a green fluorescent protein ([[green fluorescent protein|GFP]]). Calcium triggers release of the luciferin ([[coelenterazine]]) from the luciferin binding protein. The substrate is then available for [[oxidation]] by the luciferase, where it is degraded to coelenteramide with a resultant release of energy. In the absence of GFP, this energy would be released as a photon of blue light (peak emission wavelength 482&nbsp;nm). However, due to the closely associated GFP, the energy released by the luciferase is instead coupled through [[resonance energy transfer]] to the [[fluorophore]] of the GFP, and is subsequently released as a photon of green light (peak emission wavelength 510&nbsp;nm). The catalyzed reaction is:<ref name="Shimomura_1985">{{cite journal | vauthors = Shimomura O | title = Bioluminescence in the sea: photoprotein systems | journal = Symposia of the Society for Experimental Biology | volume = 39 | pages = 351–372 | year = 1985 | pmid = 2871634 }}</ref>

* [[coelenterazine]] + [[Oxygen|O<sub>2</sub>]] → [[coelenteramide]] + [[carbon dioxide|CO<sub>2</sub>]] + photon of light

==== Copepod ====

Newer luciferases have recently been identified that, unlike other luciferases, are naturally secreted molecules.  One such example is the ''Metridia'' [[coelenterazine]]-dependent luciferase (MetLuc, {{UniProt|A0A1L6CBM1}}) that is derived from the marine copepod ''[[Metridia longa]]''. The ''[[Metridia longa]]'' secreted luciferase gene encodes a 24 kDa protein containing an N-terminal secretory signal [[peptide]] of 17 [[amino acid]] residues. The sensitivity and high signal intensity of this luciferase molecule proves advantageous in many reporter studies. Some of the benefits of using a secreted reporter molecule like MetLuc is its no-lysis protocol that allows one to be able to conduct live cell assays and multiple assays on the same cell.<ref name="Huh_2009">{{cite journal | vauthors = Huh S, Lee J, Jung E, Kim SC, Kang JI, Lee J, Kim YW, Sung YK, Kang HK, Park D | title = A cell-based system for screening hair growth-promoting agents | journal = Archives of Dermatological Research | volume = 301 | issue = 5 | pages = 381–385 | date = Jun 2009 | pmid = 19277688 | doi = 10.1007/s00403-009-0931-0 | s2cid = 23916875 }}</ref>

=== Bacterial ===

Bacterial bioluminescence is seen in Photobacterium species, ''[[Vibrio fischeri]] and [[Vibrio harveyi]]''. Light emission in some [[bioluminescent bacteria]] utilizes 'antenna' such as [[lumazine]] protein to accept the energy from the primary excited state on the luciferase, resulting in an excited [[lulnazine]] [[chromophore]] which emits light that is of a shorter wavelength (more blue), while in others use a [[yellow fluorescent protein]] (YFP) with   [[flavin mononucleotide]] (FMN) as the chromophore and emits light that is red-shifted relative to that from luciferase.<ref>{{cite journal | vauthors = Baldwin TO, Christopher JA, Raushel FM, Sinclair JF, Ziegler MM, Fisher AJ, Rayment I | title = Structure of bacterial luciferase | journal = Current Opinion in Structural Biology | volume = 5 | issue = 6 | pages = 798–809 | date = Dec 1995 | pmid = 8749369 | doi = 10.1016/0959-440x(95)80014-x }}</ref>

=== Dinoflagellate ===

[[Dinoflagellate]] luciferase is a multi-[[Protein domain|domain]] [[eukaryote]] protein, consisting of an [[N-terminal]] domain, and three [[catalytic domain]]s, each of which preceded by a helical bundle domain. The [[secondary structure|structure]] of the dinoflagellate luciferase [[catalytic]] [[protein domain|domain]] has been solved.<ref name="Schultz_2005">{{cite journal | vauthors = Schultz LW, Liu L, Cegielski M, Hastings JW | title = Crystal structure of a pH-regulated luciferase catalyzing the bioluminescent oxidation of an open tetrapyrrole | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 102 | issue = 5 | pages = 1378–1383 | date = Feb 2005 | pmid = 15665092 | pmc = 547824 | doi = 10.1073/pnas.0409335102 | bibcode = 2005PNAS..102.1378S | doi-access = free }}</ref> The core part of the domain is a 10 stranded [[beta barrel]] that is [[structurally]] similar to [[lipocalin]]s and [[FABP]].<ref name="Schultz_2005" /> 
The N-terminal domain is [[conserved sequence|conserved]] between dinoflagellate luciferase and [[luciferin]] binding proteins (LBPs). It has been suggested that this region may mediate an interaction between LBP and luciferase or their association with the [[vacuolar]] membrane.<ref name="Okamoto_2001">{{cite journal | vauthors = Okamoto OK, Liu L, Robertson DL, Hastings JW | title = Members of a dinoflagellate luciferase gene family differ in synonymous substitution rates | journal = Biochemistry | volume = 40 | issue = 51 | pages = 15862–15868 | date = Dec 2001 | pmid = 11747464 | doi = 10.1021/bi011651q | citeseerx = 10.1.1.494.3563 }}</ref>
The helical bundle domain has a three [[helix bundle]] [[secondary structure|structure]] that holds four important [[histidines]] that are thought to play a role in the [[pH]] regulation of the [[enzyme]].<ref name="Schultz_2005" /> There is a large pocket in the β-barrel of the dinoflagellate luciferase at pH 8 to accommodate the [[tetrapyrrole]] substrate but there is no opening to allow the substrate to enter. Therefore, a significant conformational change must occur to provide access and space for a [[ligand]] in the active site and the source for this change is through the four N-terminal histidine residues.<ref name="Schultz_2005" /> At pH 8, it can be seen that the unprotonated histidine residues are involved in a network of [[hydrogen bonds]] at the interface of the helices in the bundle that block substrate access to the [[active site]] and disruption of this interaction by [[protonation]] (at pH 6.3) or by replacement of the histidine residues by [[alanine]] causes a large molecular motion of the bundle, separating the helices by 11Å and opening the catalytic site.<ref name="Schultz_2005" /> Logically, the histidine residues cannot be replaced by alanine in nature but this experimental replacement further confirms that the larger histidine residues block the active site. Additionally, three Gly-Gly sequences, one in the N-terminal helix and two in the helix-loop-helix motif, could serve as hinges about which the chains rotate in order to further open the pathway to the catalytic site and enlarge the active site.<ref name="Schultz_2005" />

A dinoflagellate luciferase is capable of emitting light due to its interaction with its substrate ([[luciferin]]) and the luciferin-binding protein (LBP) in the [[scintillon]] [[organelle]] found in dinoflagellates.<ref name="Schultz_2005" /> The luciferase acts in accordance with luciferin and LBP in order to emit light but each component functions at a different pH. Luciferase and its domains are not active at pH 8 but they are extremely active at the optimum pH of 6.3 whereas LBP binds luciferin at pH 8 and releases it at pH 6.3.<ref name="Schultz_2005" /> Consequently, luciferin is only released to react with an active luciferase when the scintillon is acidified to pH 6.3. Therefore, in order to lower the pH, [[voltage-gated]] channels in the scintillon [[membrane]] are opened to allow the entry of [[protons]] from a [[vacuole]] possessing an [[action potential]] produced from a mechanical stimulation.<ref name="Schultz_2005" /> Hence, it can be seen that the action potential in the vacuolar membrane leads to acidification and this in turn allows the luciferin to be released to react with luciferase in the scintillon, producing a flash of blue light.

== Reaction mechanism ==
All luciferases are classified as [[oxidoreductases]] ({{EC number|1.13.12.-}}), meaning they act on [[Electron donor|single donors]] with incorporation of [[molecular oxygen]]. Because luciferases are from many diverse [[Protein family|protein families]] that are unrelated, there is no unifying mechanism, as any mechanism depends on the luciferase and luciferin combination. However, all characterised luciferase-luciferin reactions<ref>{{Cite web |title=Everything About Luciferin and Luciferase {{!}} GoldBio |url=https://goldbio.com/articles/article/Everything-About-Luciferin-Luciferase?srsltid=AfmBOooniZZNmY5x0yA6fqZsBQgX8huANdmCT9Tq4PDyXLIl01RFcOA4 |access-date=2025-05-28 |website=goldbio.com |language=en-US}}</ref> to date have been shown to require molecular [[oxygen]] at some stage.
[[File:Active Site Firefly Luciferase.png|thumb|Firefly ''Photinus pyralis'' luciferase in the adenylate-forming conformation bound to DLSA. Key interaction observed between K529 and carbonyl oxygen of adenylate. PDB 4G36|296x296px]]Firefly ''[[Photinus pyralis]]'' luciferase in the adenylate-forming conformation bound to DLSA. Key interaction observed between K529 and carbonyl oxygen of [[adenylate]]. PDB 4G36]]

=== Firefly luciferase ===
The luciferase of ''Photinus pyralis'' catalyzes a two-step bioluminescent reaction. First is [[Adenylylation|adenylation]], a process in which D-luciferin is converted to D-luciferyl-adenylate (D-AMP) via the covalent addition of [[adenosine monophosphate]] to an amino acid side chain. Next, [[oxidative decarboxylation]] of the adenylated intermediate occurs, a necessary step for light emission. Studies have presented the first crystal structure of luciferase in its second catalytic conformation using DLSA (5′-O-[N-(dehydroluciferyl)-sulfamoyl]adenosine), a stable analog of D-AMP. The Photinus pyralis luciferase in the adenylate-forming conformation bound to DLSA illustrates conserved interactions observed in other adenylate-forming enzymes as well as key insights into the mechanism of [[bioluminescence]]. The active site is located at the interface of the N-terminal and C-terminal domains. Lys529 is the catalytic lysine for the initial adenylation reaction, interacting with the carbonyl oxygen of the ligand. A transition to the oxidation-available conformation involves a ~140° rotation of the C-terminal domain, upon which oxidation initiates formation of the dioxetanone intermediate. The decomposition of this intermediate releases visible light. Unlike D-AMP, DLSA cannot undergo oxidation, but the “locking” of the enzyme in its second catalytic conformation allowed researchers to study the oxidation-ready state of luciferase.<ref>{{cite journal | vauthors = Gulick A | title = Crystal Structure of Firefly Luciferase in a Second Catalytic Conformation Supports a Domain Alternation Mechanism | journal = Biochemistry | volume = 51 | issue = 33 | pages = 6493–6495 | date = August 1, 2012 | pmid = 22852753 | pmc = 3425952 | doi = 10.1021/bi300934s }}</ref>

{{Infobox protein family
| Symbol = Bac_luciferase
| Name = Bacterial Luciferase monooxygenase family
| width =
| Pfam = PF00296
| InterPro =IPR016048
| SMART =
| PROSITE =PDOC00397
| SCOP =1nfp
| TCDB =
| OPM family =
| OPM protein =
| PDB = {{PDB2|1brl}}, {{PDB2|1bsl}}, {{PDB2|1ezw}}, {{PDB2|1fvp}}, {{PDB2|1luc}}, {{PDB2|1m41}}, {{PDB2|1nfp}}, {{PDB2|1nqk}}, {{PDB2|1rhc}}, {{PDB2|1xkj}}
}}

=== Bacterial luciferase ===
The reaction catalyzed by bacterial luciferase is also an oxidative process:
* FMNH<sub>2</sub> + O<sub>2</sub> + RCHO → FMN + RCOOH + H<sub>2</sub>O + light

In the reaction, molecular oxygen oxidizes [[flavin mononucleotide]] and a long-chain aliphatic [[aldehyde]] to an aliphatic [[carboxylic acid]]. The reaction forms an excited hydroxyflavin intermediate, which is dehydrated to the product FMN to emit blue-green light.<ref>{{cite journal | vauthors = Fisher AJ, Thompson TB, Thoden JB, Baldwin TO, Rayment I | title = The 1.5-A resolution crystal structure of bacterial luciferase in low salt conditions | journal = The Journal of Biological Chemistry | volume = 271 | issue = 36 | pages = 21956–21968 | date = Sep 1996 | pmid = 8703001 | doi = 10.1074/jbc.271.36.21956 | doi-access = free }}</ref>
 
Nearly all of the energy input into the reaction is transformed into light. The reaction is 80%<ref>{{cite journal | vauthors = Wilson E | title = What's That Stuff? | journal = Chemical and Engineering News | volume = 77 | issue = 3 | pages = 65 | date = Jan 18, 1999 | doi = 10.1021/cen-v077n003.p065 | url = http://pubs.acs.org/cen/whatstuff/stuff/7703scit4.html | url-access = subscription }}</ref> to 90%<ref name="Knivett_2009">{{cite journal | vauthors = Knivett V | title = Lighting the way | journal = EE Times | year = 2009 | url = http://www.eetimes.com/electronics-blogs/planet-analog-designline-blog/4035489/Lighting-the-way | access-date = 2011-09-18 | archive-date = 2012-10-05 | archive-url = https://web.archive.org/web/20121005070104/http://www.eetimes.com/electronics-blogs/planet-analog-designline-blog/4035489/Lighting-the-way | url-status = dead }}</ref> efficient.  In comparison, the [[light bulb|incandescent light bulb]] only converts about 10% of its [[energy]] into light<ref>General Electric TP-110, p. 23, table.</ref> and a 150 lumen per Watt (lm/W) LED converts 20% of input energy to visible light.<ref name="Knivett_2009" />

== Applications ==
[[File:Data on circadian rhythm of gene expression in four seedlings. Two of these Arabidopsis thaliana seedlings carry a firefly luciferase reporter for transcription of the gene CCA1 and two for TOC1.png|alt=Graph with four rhythmic lines that peak in pairs, at alternating times over six, 24-hour cycles.|thumb|Graph showing timeseries data from ''in vivo'' imaging of firefly luciferase [[bioluminescence]]. [[Genetically modified organism|Transgenic]] seedlings of ''[[Arabidopsis thaliana]]'' were imaged by a cooled [[Charge-coupled device|CCD camera]] under three cycles of 12h light: 12h dark followed by 3 days of constant light. Their genomes carry firefly luciferase [[reporter gene]]s, so the signals of seedlings 61 (red) and 62 (blue) reflect transcription of the gene [[Circadian Clock Associated 1|CCA1]], while 64 (pale grey) and 65 (teal) reflect [[TOC1 (gene)|TOC1]]. The timeseries show 24-hour, [[circadian rhythm]]s of gene expression in the living plants. ]]

Luciferases can be produced in the lab through [[genetic engineering]] for a number of purposes.  Luciferase [[gene]]s can be synthesized and inserted into organisms or [[transfected]] into cells. As of 2002, [[mouse|mice]], [[silkworm]]s, and [[potato]]es are just a few of the organisms that have already been engineered to produce the protein.<ref name="Contag_2002">{{cite journal | vauthors = Contag CH, Bachmann MH | title = Advances in<!--not a mistake--> in vivo bioluminescence imaging of gene expression | journal = Annual Review of Biomedical Engineering | volume = 4 | pages = 235–260 | year = 2002 | pmid = 12117758 | doi = 10.1146/annurev.bioeng.4.111901.093336 }}</ref>

In the luciferase reaction, light is emitted when luciferase acts on the appropriate [[luciferin]] [[substrate (biochemistry)|substrate]]. Photon emission can be detected by light sensitive apparatus such as a [[luminometer]] or an [[optical microscopes|optical microscope]] with a [[CCD camera]]. This allows observation of biological processes.<ref name="urlIntroduction to Bioluminescence Assays">{{cite web | title = Introduction to Bioluminescence Assays | url = http://www.promega.com/multimedia/bioLum01.htm | publisher = Promega Corporation | archive-url = https://web.archive.org/web/20100814085557/http://www.promega.com/multimedia/bioLum01.htm | archive-date = 2010-08-14 | access-date = 2009-03-07 | url-status = dead }}</ref>  Since light excitation is not needed for luciferase bioluminescence, there is minimal [[autofluorescence]] and therefore the bioluminescent signal is virtually background-free.<ref name="Williams_1989">{{cite journal | vauthors = Williams TM, Burlein JE, Ogden S, Kricka LJ, Kant JA | title = Advantages of firefly luciferase as a reporter gene: application to the interleukin-2 gene promoter | journal = Analytical Biochemistry | volume = 176 | issue = 1 | pages = 28–32 | date = Jan 1989 | pmid = 2785354 | doi = 10.1016/0003-2697(89)90267-4 }}</ref>  Therefore, as little as 0.02&nbsp;pg can still be accurately measured using a standard [[scintillation counter]].<ref name="Nguyen_1988">{{cite journal | vauthors = Nguyen VT, Morange M, Bensaude O | title = Firefly luciferase luminescence assays using scintillation counters for quantitation in transfected mammalian cells | journal = Analytical Biochemistry | volume = 171 | issue = 2 | pages = 404–408 | date = Jun 1988 | pmid = 3407940 | doi = 10.1016/0003-2697(88)90505-2 }}</ref>

In biological research, luciferase is commonly used as a reporter to assess the [[Transcription (genetics)|transcription]]al activity in cells that are transfected with a genetic construct containing the luciferase gene under the control of a [[promoter (biology)|promoter]] of interest.<ref name="Fan_2007">{{cite journal | vauthors = Fan F, Wood KV | title = Bioluminescent assays for high-throughput screening | journal = Assay and Drug Development Technologies | volume = 5 | issue = 1 | pages = 127–136 | date = Feb 2007 | pmid = 17355205 | doi = 10.1089/adt.2006.053 }}</ref> Additionally, proluminescent molecules that are converted to luciferin upon activity of a particular enzyme can be used to detect enzyme activity in coupled or two-step luciferase assays. Such substrates have been used to detect [[caspase]] activity and [[cytochrome P450]] activity, among others.<ref name="urlIntroduction to Bioluminescence Assays"/><ref name="Fan_2007" />

Luciferase can also be used to detect the level of cellular ATP in cell viability [[assay]]s or for kinase activity assays.<ref name="Fan_2007" /><ref name="promega">{{cite journal | vauthors = Meisenheimer PL, O'Brien MA, Cali JJ | title = Luminogenic enzyme substrates: The basis for a new paradigm in assay design. | journal = Promega Notes | volume = 100 | pages = 22–26 | date = September 2008 | url = http://www.promega.com/pnotes/100/16620_22/16620_22.pdf | access-date = 2008-10-01 | archive-url = https://web.archive.org/web/20090306043643/http://www.promega.com/pnotes/100/16620_22/16620_22.pdf | archive-date = 2009-03-06 | url-status = dead }}</ref> Luciferase can act as an ATP sensor protein through [[biotinylation]]. Biotinylation will immobilize luciferase on the cell-surface by binding to a [[streptavidin]]-[[biotin]] complex. This allows luciferase to detect the efflux of ATP from the cell and will effectively display the real-time release of ATP through bioluminescence.<ref>{{cite journal | vauthors = Nakamura M, Mie M, Funabashi H, Yamamoto K, Ando J, Kobatake E | title = Cell-surface-localized ATP detection with immobilized firefly luciferase | journal = Analytical Biochemistry | volume = 352 | issue = 1 | pages = 61–67 | date = May 2006 | pmid = 16564487 | doi = 10.1016/j.ab.2006.02.019 }}</ref> Luciferase can additionally be made more sensitive for ATP detection by increasing the luminescence intensity by changing certain [[amino acid]] [[residue (chemistry)|residues]] in the sequence of the protein.<ref>{{cite journal | vauthors = Fujii H, Noda K, Asami Y, Kuroda A, Sakata M, Tokida A | title = Increase in bioluminescence intensity of firefly luciferase using genetic modification | journal = Analytical Biochemistry | volume = 366 | issue = 2 | pages = 131–136 | date = Jul 2007 | pmid = 17540326 | doi = 10.1016/j.ab.2007.04.018 }}</ref>

Whole organism imaging (referred to as ''in vivo'' when intact or, otherwise called ''ex vivo'' imaging for example of living but explanted tissue) is a powerful technique for studying cell populations in live plants or animals, such as mice.<ref name="Greer_2002">{{cite journal | vauthors = Greer LF, Szalay AA | title = Imaging of light emission from the expression of luciferases in living cells and organisms: a review | journal = Luminescence | volume = 17 | issue = 1 | pages = 43–74 | year = 2002 | pmid = 11816060 | doi = 10.1002/bio.676 | doi-access = free }}</ref> Different types of cells (e.g. bone marrow stem cells, T-cells) can be engineered to express a luciferase allowing their non-invasive visualization inside a live animal using a sensitive charge-couple device camera ([[CCD camera]]).This technique has been used to follow tumorigenesis and response of tumors to treatment in animal models.<ref name="Lyons_2003">{{cite journal | vauthors = Lyons SK, Meuwissen R, Krimpenfort P, Berns A | title = The generation of a conditional reporter that enables bioluminescence imaging of Cre/loxP-dependent tumorigenesis in mice | journal = Cancer Research | volume = 63 | issue = 21 | pages = 7042–7046 | date = Nov 2003 | pmid = 14612492 | url = http://cancerres.aacrjournals.org/cgi/content/abstract/63/21/7042 }}</ref><ref name="Becher_2006">{{cite journal | vauthors = Becher OJ, Holland EC | title = Genetically engineered models have advantages over xenografts for preclinical studies | journal = Cancer Research | volume = 66 | issue = 7 | pages = 3355–8, discussion 3358–9 | date = Apr 2006 | pmid = 16585152 | doi = 10.1158/0008-5472.CAN-05-3827 | doi-access = free }}</ref> However, environmental factors and therapeutic interferences may cause some discrepancies between tumor burden and bioluminescence intensity in relation to changes in proliferative activity. The intensity of the signal measured by in vivo imaging may depend on various factors, such as <small>D</small>-luciferin absorption through the peritoneum, blood flow, cell membrane permeability, availability of co-factors, [[intracellular pH]] and transparency of overlying tissue, in addition to the amount of luciferase.<ref>{{cite journal | vauthors = Inoue Y, Tojo A, Sekine R, Soda Y, Kobayashi S, Nomura A, Izawa K, Kitamura T, Okubo T, Ohtomo K | title = In vitro validation of bioluminescent monitoring of disease progression and therapeutic response in leukaemia model animals | journal = European Journal of Nuclear Medicine and Molecular Imaging | volume = 33 | issue = 5 | pages = 557–565 | date = May 2006 | pmid = 16501974 | doi = 10.1007/s00259-005-0048-4 | s2cid = 40630078 }}</ref>

Luciferase is a heat-sensitive protein that is used in studies on [[protein denaturation]], testing the protective capacities of [[heat shock protein]]s. The opportunities for using luciferase continue to expand.<ref name="Massoud_2007">{{cite journal | vauthors = Massoud TF, Paulmurugan R, De A, Ray P, Gambhir SS | title = Reporter gene imaging of protein-protein interactions in living subjects | journal = Current Opinion in Biotechnology | volume = 18 | issue = 1 | pages = 31–37 | date = Feb 2007 | pmid = 17254764 | pmc = 4141564 | doi = 10.1016/j.copbio.2007.01.007 }}</ref>

== See also ==
* [[Quorum sensing]]
{{clear}}

== References ==
{{Reflist|33em}}

== Further reading ==
{{refbegin|33em}}
* {{cite web | vauthors = Goodsell D | title = Luciferase | url = http://www.rcsb.org/pdb/101/motm.do?momID=78 | work = Molecule of the Month | publisher = Protein Data Bank | access-date = 2013-01-15 | archive-date = 2015-10-19 | archive-url = https://web.archive.org/web/20151019150930/http://www.rcsb.org/pdb/101/motm.do?momID=78 | url-status = dead }}
* {{cite web | vauthors = Haddock SH, McDougall CM, Case JF | title = Luciferin types | work = The Bioluminescence Web Page | url = http://www.lifesci.ucsb.edu/~biolum/chem/detail1.html | orig-date = 1997 | date = 2011 | publisher = University of California, Santa Barbara | access-date = 2009-03-07 }}
* {{cite web | vauthors = Trimmer B, Zayas R, Qazi S, Lewis S, Michel T, Dudzinski D, Aprille J, Lagace C | title = Firefly flashes and Nitric Oxide | date = 2001-06-28 | url = http://ase.tufts.edu/biology/Firefly/ | archive-url = https://web.archive.org/web/20120128044100/http://ase.tufts.edu/biology/Firefly/ | archive-date = 28 January 2012 | publisher = Tufts University | access-date = 2008-10-02 }}
* {{cite web | title = Bioluminescence Reporters Guide | url = http://www.promega.com/paguide/chap8.htm | work = Protocols and applications | publisher = Promega Corporation | archive-url = https://web.archive.org/web/20100808140556/http://www.promega.com/paguide/chap8.htm | archive-date = 2010-08-08 | access-date = 2009-03-07 | url-status = dead }}
* {{cite encyclopedia | title = Luciferin types | url = http://www.iscid.org/encyclopedia/Luciferase | encyclopedia = ISCID Encyclopedia of Science and Philosophy | publisher = International Society for Complexity, Information, and Design (ISCID)
 | archive-url = https://web.archive.org/web/20120921155432/http://www.iscid.org/encyclopedia/Luciferase | archive-date = 2012-09-21 | access-date = 2010-04-20 | url-status = dead }}
{{refend}}

== External links ==
{{refbegin}}
* {{Commons category-inline}}
* {{PDBe-KB2|P08659|Luciferin 4-monooxygenase}}
{{refend}}

{{Monooxygenases}}
{{Enzymes}}
{{Portal bar|Biology|border=no}}

{{InterPro content|IPR018804}}
{{InterPro content|IPR007959}}
{{InterPro content|IPR018475}}

[[Category:Protein domains]]
[[Category:Oxidoreductases]]
[[Category:Bioluminescence]]
[[Category:Enzymes of known structure]]