Difference between revisions of "Part:BBa K3137005"

Latest revision as of 01:59, 22 October 2019

gntR

One of the most abundant and widely distributed groups of Helix-turn-helix (HTH) transcription factors is the metabolite-responsive GntR family of regulators. These proteins contain a DNA-binding HTH domain at the N-terminus of the protein and an effector-binding and/or oligomerisation domain at the C-terminus, where upon on binding an effector molecule, a conformational change occurs in the protein which influences the DNA-binding properties of the regulator resulting in repression or activation of transcription.

Usage and Biology

To get more efficient phage resistant parts, we used the ARTP(Atmospheric and room temperature plasma) mutagenesis system and Comparative Genomics to obtain 4 mutant bacterial and 4 key mutant sites (nuoE, yhjH, rzpD And gntR).Resistance tests on these key sites were performed respectively(Fig 1&2).

Figure 1. Resistance test for T4 phage of NuoE, YhjH, RzpD and GntR

Figure 2. Resistance test data for T4 phage of NuoE, YhjH, RzpD and GntR

Since it is not possible to directly see from the figure which component has the least influence on the growth of the bacteria, we use the Grey Relation Analysis (GRA) method to analyze the growth curve (Fig 3) of the bacteria connecting the various components.

Figure 3. Effect of different anti-phage parts on the growth of E. coli BL21

We used the Entropy Weight Method (EWM) to determine the weight of each growth point to select the most similarly modified strain (the highest correlation), which is the component that has the least impact on bacterial growth. At the same time, after consulting the industry experts, we revised the weights according to the experts' recommendations to evaluate the components again, making them more in line with the real situation of production, that is, the most suitable components for industrial production. Using GRA's two evaluations of the four components at different weights, we selected the component gntR.(Fig 4)

Figure 4. EWM weight & advice weight and the results of GRA for selecting most suitable strain

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 425
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

@@ Line 3: / Line 3: @@
 <partinfo>BBa_K3137005 short</partinfo>
-The Gluconate repressor gntR, is a transcription factor that negatively regulates the operon involved in the catabolism of d-gluconate via the Entner-Doudoroff pathway and represses genes involved in the different systems related to d-gluconate uptake: gluconate Ⅰ and gluconate Ⅱ.
+One of the most abundant and widely distributed groups of Helix-turn-helix (HTH) transcription factors is the metabolite-responsive GntR family of regulators. These proteins contain a DNA-binding HTH domain at the N-terminus of the protein and an effector-binding and/or oligomerisation domain at the C-terminus, where upon on binding an effector molecule, a conformational change occurs in the protein which influences the DNA-binding properties of the regulator resulting in repression or activation of transcription.
+===Usage and Biology===
+To get more efficient phage resistant parts, we used the ARTP(Atmospheric and room temperature plasma) mutagenesis system and Comparative Genomics to obtain 4 mutant bacterial and 4 key mutant sites (<i>nuoE</i>, <i>yhjH</i>, <i>rzpD</i> And <i>gntR</i>).Resistance tests on these key sites were performed respectively(Fig 1&2).
+[[Image:sapp1.jpg|500px|center|thumb|Figure 1. Resistance test for T4 phage of NuoE, YhjH, RzpD and GntR]]
+[[Image:Sapp5.png|500px|center|thumb|Figure 2. Resistance test data for T4 phage of NuoE, YhjH, RzpD and GntR]]
+Since it is not possible to directly see from the figure which component has the least influence on the growth of the bacteria, we use the Grey Relation Analysis (GRA) method to analyze the growth curve (Fig 3) of the bacteria connecting the various components.
+[[Image:sapp2.jpg|500px|center|thumb|Figure 3. Effect of different anti-phage parts on the growth of <i>E. coli</i> BL21]]
+We used the Entropy Weight Method (EWM) to determine the weight of each growth point to select the most similarly modified strain (the highest correlation), which is the component that has the least impact on bacterial growth. At the same time, after consulting the industry experts, we revised the weights according to the experts' recommendations to evaluate the components again, making them more in line with the real situation of production, that is, the most suitable components for industrial production. Using GRA's two evaluations of the four components at different weights, we selected the component <i>gntR</i>.(Fig 4)
+[[Image:sapp3.png|800px|center|thumb|Figure 4. EWM weight & advice weight and the results of GRA for selecting most suitable strain]]