Difference between revisions of "Part:BBa K4274031"
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<partinfo>BBa_K4274031 short</partinfo> | <partinfo>BBa_K4274031 short</partinfo> | ||
| − | ptac-RiboJ-B0034-ClSS_S533A-B0034-ERG20_F96W-B0015 was used to complete the pathway of IPP ⇄DMAPP → GPP → FPP → alpha-santalene. This part expresses the necessary enzymes (C. lansium santalene synthase and ERG20) participating in the formation of alpha-santalene. This composite part utilizes the promoter ptac-RiboJ (BBa_K3552015), which has an optimized productivity and is IPTG-inducible to initiate the later sequence. ptac-RiboJ is followed by the ribosome binding site (BBa_B0034), derived from the Elowitz repressilator. Then, we coded in the | + | ptac-RiboJ-B0034-ClSS_S533A-B0034-ERG20_F96W-B0015 was used to complete the pathway of IPP ⇄DMAPP → GPP → FPP → alpha-santalene. This part expresses the necessary enzymes (<i>C. lansium</i> santalene synthase and ERG20) participating in the formation of alpha-santalene. This composite part utilizes the promoter ptac-RiboJ (BBa_K3552015), which has an optimized productivity and is IPTG-inducible to initiate the later sequence. ptac-RiboJ is followed by the ribosome binding site (BBa_B0034), derived from the Elowitz repressilator. Then, we coded in the <i>Cl</i>SS_S533A(BBa_K4274000), which mutated its 533th amino acid from serine to alanine, reaching the optimization of its effectiveness. <i>C. lansium</i> santalene synthase plays a crucial role to assist the transformation from FPP (farnesyl diphosphate) to alpha-santalene (the precursor of santalol). Then, after an identical ribosome binding site as before (BBa_B0034), we inserted the sequence for ERG20_F96W, which can catalyze the formation of FPP (farnesyl diphosphate). Compared with ERG20（BBa_K849001），ERG20_F96W’s 96th amino acid being mutated from phenylalanine to tryptophan turns out to significantly improve the yield of a-santalene. This part ends with the double terminator (BBa_B0015). |
| − | We have created a parts collection for all parts optimizing the production of alpha-santalene. This collection includes the series to optimize the activity of farnesyl pyrophosphate synthase, ERG20_F96W (Part: BBa_K4274002) and ptac-RiboJ-B0034- | + | We have created a parts collection for all parts optimizing the production of alpha-santalene. This collection includes the series to optimize the activity of farnesyl pyrophosphate synthase, ERG20_F96W (Part: BBa_K4274002) and ptac-RiboJ-B0034-<i>Cl</i>SS_S533A-B0034-ERG20-B0015 (Part: BBa_K4274030). A series conducting the secretion of <i>Cl</i>SS and ERG20, consisting of CmR29M2-<i>Cl</i>SS_S533A (Part: BBa_K4274001) and ptac-RiboJ-B0034-CmR29M2-ClSS_S533A -B0034-ERG20_F96W-B0015 (Part: BBa_K4274032). However, the comparative results revealed that this modification does not optimize the production of alpha-santalene. Finally, the series of parts expressing fusion protein <i>Cl</i>SS_S533A-FL-ERG20_F96W (Part: BBa_K4274033), and BBa_K4274003. |
We aim to assist future teams in their journey, providing a collection that optimizes the production of santalene. | We aim to assist future teams in their journey, providing a collection that optimizes the production of santalene. | ||
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==Usage and Biology== | ==Usage and Biology== | ||
| − | + | <i>Cl</i>SS_S533A refers to the mutated and optimized <i>C. lansium</i> santalene synthase, an enzyme that assists the transformation from FPP (farnesyl diphosphate) to alpha-santalene. ERG20_F96W is a mutated synthase that catalyzes the formation of FPP in the pathway that produces santalene. This composite part can express both optimized enzymes for the higher production of a-santalene. | |
==Source== | ==Source== | ||
| − | + | <i>Cl</i>SS_S533A (Part: BBa_K4274000) is from <i>Clausena lansium</i>, and ERG20_F96W (Part: BBa_K849001) is from <i>S. cerevisiae</i>. | |
==Sequence and features== | ==Sequence and features== | ||
Revision as of 10:47, 12 October 2022
ptac-RiboJ-B0034-ClSS_S533A-B0034-ERG20_F96W-B0015
ptac-RiboJ-B0034-ClSS_S533A-B0034-ERG20_F96W-B0015 was used to complete the pathway of IPP ⇄DMAPP → GPP → FPP → alpha-santalene. This part expresses the necessary enzymes (C. lansium santalene synthase and ERG20) participating in the formation of alpha-santalene. This composite part utilizes the promoter ptac-RiboJ (BBa_K3552015), which has an optimized productivity and is IPTG-inducible to initiate the later sequence. ptac-RiboJ is followed by the ribosome binding site (BBa_B0034), derived from the Elowitz repressilator. Then, we coded in the ClSS_S533A(BBa_K4274000), which mutated its 533th amino acid from serine to alanine, reaching the optimization of its effectiveness. C. lansium santalene synthase plays a crucial role to assist the transformation from FPP (farnesyl diphosphate) to alpha-santalene (the precursor of santalol). Then, after an identical ribosome binding site as before (BBa_B0034), we inserted the sequence for ERG20_F96W, which can catalyze the formation of FPP (farnesyl diphosphate). Compared with ERG20（BBa_K849001），ERG20_F96W’s 96th amino acid being mutated from phenylalanine to tryptophan turns out to significantly improve the yield of a-santalene. This part ends with the double terminator (BBa_B0015).
We have created a parts collection for all parts optimizing the production of alpha-santalene. This collection includes the series to optimize the activity of farnesyl pyrophosphate synthase, ERG20_F96W (Part: BBa_K4274002) and ptac-RiboJ-B0034-ClSS_S533A-B0034-ERG20-B0015 (Part: BBa_K4274030). A series conducting the secretion of ClSS and ERG20, consisting of CmR29M2-ClSS_S533A (Part: BBa_K4274001) and ptac-RiboJ-B0034-CmR29M2-ClSS_S533A -B0034-ERG20_F96W-B0015 (Part: BBa_K4274032). However, the comparative results revealed that this modification does not optimize the production of alpha-santalene. Finally, the series of parts expressing fusion protein ClSS_S533A-FL-ERG20_F96W (Part: BBa_K4274033), and BBa_K4274003.
We aim to assist future teams in their journey, providing a collection that optimizes the production of santalene.
Usage and Biology
ClSS_S533A refers to the mutated and optimized C. lansium santalene synthase, an enzyme that assists the transformation from FPP (farnesyl diphosphate) to alpha-santalene. ERG20_F96W is a mutated synthase that catalyzes the formation of FPP in the pathway that produces santalene. This composite part can express both optimized enzymes for the higher production of a-santalene.
Source
ClSS_S533A (Part: BBa_K4274000) is from Clausena lansium, and ERG20_F96W (Part: BBa_K849001) is from S. cerevisiae.
Sequence and features
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 1541
- 1000COMPATIBLE WITH RFC[1000]