Difference between revisions of "Part:BBa K325219"
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| + | <partinfo>BBa_K325219 short</partinfo> | ||
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D-Luciferin has to be added to obtain light output. | D-Luciferin has to be added to obtain light output. | ||
| + | The light-emitting reaction involves the conversion of D-Luciferin into oxyluciferin. This compound competes with D-Luciferin for the lucifearase's binding site, causing strong inhibition of enzyme activity. LRE removes oxyluciferin from the system by converting it into 2-cyano- 6-hydroxybenzothiazole (CHBT). This compound is non-enzymatically converted into D-Luciferin in the presence of D-cysteine. It has been proposed that in the natural system, L-cysteine is used to produce L-Luciferin, which then isomerises into D-Luciferin, but this could not be reproduced by the 2010 Cambridge iGEM team. | ||
| − | + | Light output can also be achieved by addition of CHBT and D-cysteine instead of D-Luciferin, but D-cysteine might have detrimental effects on cell growth and physiology. | |
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| − | + | ==Pictures== | |
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| − | + | [[Image:300px-Cambridge-Wed.jpg|thumb|569px|center|'''Figure 1 - E.Coli (Invitrogen TOP 10) cells transformed with [https://parts.igem.org/Part:BBa_K325909 BBa K325909] (blue light bulb) and [https://parts.igem.org/Part:BBa_K325219 BBa 325219] (red light bulb) ''']] | |
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| − | + | <sup>1</sup>Measured by the [http://2010.igem.org/Team:Cambridge Cambridge iGEM team 2010] | |
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| − | <sup>1</sup>Measured by | + | |
<div style="padding: 00px; width: 680px"> | <div style="padding: 00px; width: 680px"> | ||
| + | ==Derivative parts== | ||
| + | [[Image:Cropbow.jpg|thumb|569px|center|The E.glowli team used site-directed mutagenesis to create a series of colour mutants from this BioBrick]] | ||
'''Compatibility'''<br> | '''Compatibility'''<br> | ||
| − | [https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=cell ''Chassis:''] Device has been shown to work in '' | + | [https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=cell ''Chassis:''] Device has been shown to work in ''Top 10 (Invitrogen)''<br> |
| − | [[Plasmid backbones|''Plasmids:'']] Device has been shown to work on ''<partinfo> | + | [[Plasmid backbones|''Plasmids:'']] Device has been shown to work on ''<partinfo>pSB1C3</partinfo>'' <br> |
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</div> | </div> | ||
'''References'''<br> | '''References'''<br> | ||
| − | [http://www.ncbi.nlm.nih.gov/pubmed/18949818 '''[1 | + | [http://www.ncbi.nlm.nih.gov/pubmed/18949818 '''[1]:'''] S.M. Marques and J.C.G. Esteves da Silva, (2009) Firefly Bioluminescence: A Mechanistic Approach of Luciferase Catalyzed Reactions,''Life'' '''61''', 6-17. |
</div> | </div> | ||
| + | [http://www.nature.com/nature/journal/v440/n7082/abs/nature04542.html '''[2]:'''] T. Nakatsu ''et al.'' (2006) Structural Basis for the spectral difference in luciferase bioluminescence, ''Nature'' '''440'''(16), 372-376. | ||
| + | [http://www.ncbi.nlm.nih.gov/pubmed/11457857 '''[3]:'''] K. Gomi and N. Kajiyama, (2001) Oxyluciferin, a Luminescence Product of Firefly Luciferase, Is Enzymatically Regenerated into Luciferin, ''The Journal of Biological Chemistry'', '''276'''(39), 36508-36513. | ||
Latest revision as of 10:47, 13 June 2021
Red Firefly Luciferase and LRE (under pBAD)
L. Cruciata
(E. coli optimised)
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Description
This part generates a red-mutant of the luciferase from the Japanese firefly (L.cruciata) as well as the luciferin regenerating enzyme (LRE). It is under the control of an Arabinose induced promoter.
D-Luciferin has to be added to obtain light output.
The light-emitting reaction involves the conversion of D-Luciferin into oxyluciferin. This compound competes with D-Luciferin for the lucifearase's binding site, causing strong inhibition of enzyme activity. LRE removes oxyluciferin from the system by converting it into 2-cyano- 6-hydroxybenzothiazole (CHBT). This compound is non-enzymatically converted into D-Luciferin in the presence of D-cysteine. It has been proposed that in the natural system, L-cysteine is used to produce L-Luciferin, which then isomerises into D-Luciferin, but this could not be reproduced by the 2010 Cambridge iGEM team.
Light output can also be achieved by addition of CHBT and D-cysteine instead of D-Luciferin, but D-cysteine might have detrimental effects on cell growth and physiology.
Pictures
1Measured by the [http://2010.igem.org/Team:Cambridge Cambridge iGEM team 2010]
Derivative parts
Compatibility
Chassis: Device has been shown to work in Top 10 (Invitrogen)
Plasmids: Device has been shown to work on pSB1C3
References
[http://www.ncbi.nlm.nih.gov/pubmed/18949818 [1]:] S.M. Marques and J.C.G. Esteves da Silva, (2009) Firefly Bioluminescence: A Mechanistic Approach of Luciferase Catalyzed Reactions,Life 61, 6-17.
[http://www.nature.com/nature/journal/v440/n7082/abs/nature04542.html [2]:] T. Nakatsu et al. (2006) Structural Basis for the spectral difference in luciferase bioluminescence, Nature 440(16), 372-376.
[http://www.ncbi.nlm.nih.gov/pubmed/11457857 [3]:] K. Gomi and N. Kajiyama, (2001) Oxyluciferin, a Luminescence Product of Firefly Luciferase, Is Enzymatically Regenerated into Luciferin, The Journal of Biological Chemistry, 276(39), 36508-36513.