|
|
| (60 intermediate revisions by 3 users not shown) |
| Line 29: |
Line 29: |
| | <h3>The Lux System</h3> | | <h3>The Lux System</h3> |
| | <p> | | <p> |
| − | The bacterial lux system is principally composed of five genes - LuxA, LuxB, LuxC, LuxD, and LuxE. | + | The bacterial Lux system is principally composed of five genes - <i>lux</i>A, <i>lux</i>B, <i>lux</i>C, <i>lux</i>D, and <i>lux</i>E. |
| | + | <center><a href="https://static.igem.org/mediawiki/2015/9/9f/UCSD_generallux.png" data-lightbox="lab2" data-title="In their host bacteria, the <i>lux</i> genes are not sequentially ordered, with A and B flanked by C, D, and E."><img width="700" src="https://static.igem.org/mediawiki/2015/9/9f/UCSD_generallux.png"></a></center> |
| | + | <div class="caption"><center><i>In their host bacteria, the </i>lux<i> genes are not sequentially ordered, with A and B flanked by C, D, and E.</i></center></div> |
| | + | <p> |
| | + | The proteins coded by these genes associate to form two enzymatic complexes, with <i>lux</i>A and <i>lux</i>B coding for luciferase itself and <i>lux</i>CDE coding for a fatty acid reductase complex. This reductase complex serves to provide the substrates (fatty aldehydes) for the system’s bioluminescent reaction<sup>1</sup>. Catalyzed by luciferase, these aldehydes react with FMNH<sub>2</sub> and oxygen, emitting a photon and producing a fatty acid, FMN, and water<sup>1</sup>. FMNH<sub>2</sub> is then regenerated by a flavin reductase<sup>1</sup>. |
| | <br><br> | | <br><br> |
| − | <center><a href="https://static.igem.org/mediawiki/2015/ 9/ 9f/ UCSD_generallux.png" data-lightbox=" lab2" data-title=" In their host bacteria, the lux genes are not sequentially ordered, with A and B flanked by C, D, and E."><img width=" 700" src="https://static.igem.org/mediawiki/2015/ 9/ 9f/ UCSD_generallux.png"></a></center> | + | <center><a href="https://static.igem.org/mediawiki/2015/ 1/ 1a/ UCSD_luciferase.png" data-lightbox=" lab3" data-title=" The alpha subunit of luciferase drives enzymatic activity, while the beta subunit offers structural support and stabilizes the alpha subunit as it undergoes conformational changes<sup>2</sup>."><img width=" 300" src="https://static.igem.org/mediawiki/2015/ 1/ 1a/ UCSD_luciferase.png"></a></center> |
| | <br> | | <br> |
| − | <center>In their host bacteria, the lux genes are not sequentially ordered, with A and B flanked by C, D, and E.</center> | + | <div class="caption"><center><i>The alpha subunit (blue) of luciferase drives enzymatic activity, while the beta subunit (red) offers structural support and stabilizes the alpha subunit as it undergoes conformational changes<sup>2</sup>.</i></center></div> |
| − | <br><br> | + | |
| | | | |
| − | The proteins coded by these genes associate to form two enzymatic complexes, with LuxA+LuxB coding for luciferase itself and LuxCDE coding for a fatty acid reductase complex. This reductase complex serves to provide the substrates (fatty aldehydes) for the system’s bioluminescent reaction<sup>1</sup>. Catalyzed by luciferase, these aldehydes react with FMNH2 and oxygen, emitting a photon and producing a fatty acid, FMN, and water<sup>1</sup>. FMNH2 is then regenerated by a flavin reductase<sup>1</sup>.
| + | <p> |
| − | <br><br>
| + | To allow for continuous light output, the fatty acid reductase complex recycles the fatty acid product in the luciferase-catalyzed reaction and converts it to a the substrate aldehyde. The genes <i>lux</i>C, <i>lux</i>D, and <i>lux</i>E code for a reductase, transferase, and synthetase, respectively. Together, they associate into a complex consisting of four of each enzyme<sup>1</sup>. |
| − | [Complex diagram w/ elaborating caption: The alpha subunit of luciferase drives enzymatic activity, while the beta subunit offers structural support and stabilizes the alpha subunit as it undergoes conformational changes<sup>2</sup>.]
| + | |
| − | <br><br>
| + | <center><a href="https://static.igem.org/mediawiki/2015/1/10/Fattyacidreductase.png" data-lightbox="lab3" data-title="Substrates are recruited by the transferase and moved to a synthetase-reductase complex. These associated enzymes produce a microenvironment that stabilizes reaction intermediates."><img width="700" src="https://static.igem.org/mediawiki/2015/1/10/Fattyacidreductase.png"></a></center> |
| − | To allow for continuous light output, the fatty acid reductase complex recycles the fatty acid product in the luciferase-catalyzed reaction and converts it to a the substrate aldehyde. LuxC, LuxD, and LuxE code for a reductase, transferase, and synthetase, respectively. Together, they associate into a complex consisting of four of each enzyme<sup>1</sup>. | + | |
| − | <br><br>
| + | |
| − | </p>
| + | |
| − | <center><a href="https://static.igem.org/mediawiki/2015/1/10/Fattyacidreductase.png" data-lightbox="lab3" data-title="Substrates are recruited by the transferase and moved to a synthetase-reductase complex. These associated enzymes produce a microenvironment that stabilizes reaction intermediates."><img width="700" src="https://static.igem.org/mediawiki/2015/1/10/Fattyacidreductase.png"></a> | + | |
| | <br> | | <br> |
| − | Substrates are recruited by the transferase and moved to a synthetase-reductase complex. These associated enzymes produce a microenvironment that stabilizes reaction intermediates.</center> | + | <div class="caption"><center><i>Substrates are recruited by the transferase and moved to a synthetase-reductase complex. These associated enzymes produce a microenvironment that stabilizes reaction intermediates.</i></center></div> |
| − | | + | |
| | <p> | | <p> |
| | Because the luminescent yield of the system is based on the function of these two enzymatic complexes, modifying the protein levels of each enzyme allows us to control the system’s output. | | Because the luminescent yield of the system is based on the function of these two enzymatic complexes, modifying the protein levels of each enzyme allows us to control the system’s output. |
| Line 53: |
Line 51: |
| | <p> | | <p> |
| | [1] Meighen, Edward A. "Enzymes and genes from the lux operons of bioluminescent bacteria." Annual Reviews in Microbiology 42.1 (1988): 151-176. <br> | | [1] Meighen, Edward A. "Enzymes and genes from the lux operons of bioluminescent bacteria." Annual Reviews in Microbiology 42.1 (1988): 151-176. <br> |
| − | [2] 4) Meighen, E. A., Nicoli M. Z., and Hastings, J. W. “Functional Differences of the Nonidentical Subunits of Bacterial Luciferase, Properties of Hybrids of Native and Chemically Modified Bacterial Luciferase.” Biochemistry (2003) | + | [2] Meighen, E. A., Nicoli M. Z., and Hastings, J. W. “Functional Differences of the Nonidentical Subunits of Bacterial Luciferase, Properties of Hybrids of Native and Chemically Modified Bacterial Luciferase.” Biochemistry (2003) |
| | </p> | | </p> |
| | </div> | | </div> |
