Difference between revisions of "Part:BBa K3846357"

Latest revision as of 11:36, 20 October 2021

Costunolide production casette

Mevalonate pathway

The synthesis of sesquiterpenes, our main focus of this project, relies on the availability of common building blocks like farnesyl diphosphate (FPP). It has been shown for different organisms that the amount of FPP can be increased by overexpressing proteins of the mevalonat (MevT) and the MBIS pathway which we organised to be expressed into operon structures for our project. Both operons are IPTG-inducible and use lacUV5 promoter and rrnB T1 and T2 terminator sequence.

MevT operon: This operon contains the acetoacetyl-CoA synthase (ackA), HMG-CoA synthases (HMG), and an N-terminally truncated version of HMG-CoA reductase (tHMGR).

MBIS operon: This operon contains mevalonate kinase (MK), phosphomevalonate kinase (PMK), phosphomevalonate decarboxylase (PMD), isopentenyl diphosphate isomerase (idi), and farnesyl diphosphate synthase (ispA).

By combining both of these operons in a single composite part and further adding synthases, it is possible to synthesize different bioactive compounds.

Parthenolide

Parthenolide is an interesting sesquiterpene lactone which exhibits a broad spectrum of biological activities (anti-cancer drug and migraine prophylaxis) and can be isolated from Tanacetum parthenium . Parthenolide is derived from the terpene germacrene A, which itself is a cyclization product of FPP by the germacrene A synthase. The biosynthesis of parthenolide occurs through several CYP450 enzymes (germacrene A hydroxylase, costunolide synthase and parthenolide synthase). We organised the necessary enzyme of this pathway also in an operon structure under the control of an IPTG-inducible T7 promoter.

BBa_K3846357

This part combines the MevT, MBIS and costunolide operons with the expression of the CYP P450-NAPH-reductase ATR2 and enables E. coli to produce costunolide.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 9575
Illegal NheI site found at 9579
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

@@ Line 3: / Line 3: @@
 <partinfo>BBa_K3846357 short</partinfo>
-Costunolide production casette
+===Mevalonate pathway===
+The synthesis of sesquiterpenes, our main focus of this project, relies on the availability of common building blocks like farnesyl diphosphate (FPP). It has been shown for different organisms that the amount of FPP can be increased by overexpressing proteins of the mevalonat (MevT) and the MBIS pathway which we organised to be expressed into operon structures for our project.
+Both operons are IPTG-inducible and use lacUV5 promoter and rrnB T1 and T2 terminator sequence.
+<b>MevT operon</b>:
+This operon contains the acetoacetyl-CoA synthase (ackA), HMG-CoA synthases (HMG), and an N-terminally truncated version of HMG-CoA reductase (tHMGR).
+<b>MBIS operon</b>:
+This operon contains mevalonate kinase (MK), phosphomevalonate kinase (PMK), phosphomevalonate decarboxylase (PMD), isopentenyl diphosphate isomerase (idi), and farnesyl diphosphate synthase (ispA).
+By combining both of these operons in a single composite part and further adding synthases, it is possible to synthesize different bioactive compounds.
+===Parthenolide===
+Parthenolide is an interesting sesquiterpene lactone which exhibits a broad spectrum of biological activities (anti-cancer drug and migraine prophylaxis) and can be isolated from <i>Tanacetum parthenium </i>. Parthenolide is derived from the terpene germacrene A, which itself is a cyclization product of FPP by the germacrene A synthase. The biosynthesis of parthenolide occurs through several CYP450 enzymes (germacrene A hydroxylase, costunolide synthase and parthenolide synthase).
+We organised the necessary enzyme of this pathway also in an operon structure under the control of an IPTG-inducible T7 promoter.
+===BBa_K3846357===
+This part combines the MevT, MBIS and costunolide operons with the expression of the CYP P450-NAPH-reductase ATR2 and enables <i>E. coli</i> to produce costunolide.
 <!-- Add more about the biology of this part here