Difference between revisions of "Part:BBa K4621000"

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The Ectoine is then activated by ectoine hydroxylase, which is encoded by the ectD gene, to generate 5-Hydroxyectoine. The synthesis of 5-Hydroxyectoine from L-Aspartate occurs naturally in some moderately halophilic microorganisms, as shown in Figure 1.
 
The Ectoine is then activated by ectoine hydroxylase, which is encoded by the ectD gene, to generate 5-Hydroxyectoine. The synthesis of 5-Hydroxyectoine from L-Aspartate occurs naturally in some moderately halophilic microorganisms, as shown in Figure 1.
  
https://static.igem.wiki/teams/4621/wiki/parts/gel-electrophoresis-results.png
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[[Image:pathways-for-5-hydroxyectoine-synthesis.jpg|thumbnail|750px|center|'''Figure 1:'''  
 
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[[Image:https://static.igem.wiki/teams/4621/wiki/parts/pathways-for-5-hydroxyectoine-synthesis.jpg|thumbnail|750px|center|'''Figure 1:'''  
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Pathways for 5-hydroxyectoine synthesis]]
 
Pathways for 5-hydroxyectoine synthesis]]
  

Revision as of 05:25, 2 October 2023


EctOperon is a cluster of genes encoding four enzymes that synthesize 5-Hydroxyectoine


Usage, Biology and Characterization:

EctOperon contains ectA, ectB, ectC, and ectD.

ectB is the coding sequence of L-2, 4-diaminobutyrate --2-oxoglutarate aminotransferase, L-Aspartate 4-semialdehyde can be converted to L-2,4-diaminobutanoate. ectA is the coding sequence for L-2,4-diaminobutyric acid acetyltransferase, L-2, 4-diaminobutanoate can be converted to N4-acetyl-L-2,4-diaminobutyrate. ectC is the coding sequence of L-ectoine synthase, which can convert N4-acetyl-L-2,4-diaminobutyrate to L-Ectoine. These three genes encode enzymes Ectoine: (4S)-2-methyl-1,4,5, 6-tetrahydropyrim idine-4-carboxylic acid, located on the same operon, forming the ectABC gene cluster. The Ectoine is then activated by ectoine hydroxylase, which is encoded by the ectD gene, to generate 5-Hydroxyectoine. The synthesis of 5-Hydroxyectoine from L-Aspartate occurs naturally in some moderately halophilic microorganisms, as shown in Figure 1.

File:Pathways-for-5-hydroxyectoine-synthesis.jpg
Figure 1: Pathways for 5-hydroxyectoine synthesis

Streptomyces SCUT-3 was modified to overexpressed EctOperon. They were then seeded in a 40 mL LB medium system. After fermentation at 40 °C and 220 rpm for 72h, the product 5-Hydroxyectoine was determined as follows.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 3227
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 3227
    Illegal NotI site found at 571
    Illegal NotI site found at 1456
    Illegal NotI site found at 1774
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 753
    Illegal XhoI site found at 927
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 3227
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 3227
    Illegal NgoMIV site found at 788
    Illegal NgoMIV site found at 1194
    Illegal NgoMIV site found at 1200
    Illegal NgoMIV site found at 1491
    Illegal NgoMIV site found at 2540
    Illegal NgoMIV site found at 2732
    Illegal NgoMIV site found at 3242
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 376
    Illegal BsaI.rc site found at 556
    Illegal BsaI.rc site found at 1338
    Illegal BsaI.rc site found at 1866
    Illegal BsaI.rc site found at 2121
    Illegal BsaI.rc site found at 2211
    Illegal BsaI.rc site found at 2286
    Illegal SapI site found at 414