Difference between revisions of "Part:BBa K3712006"
 
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<partinfo>BBa_K3712006 short</partinfo>
 
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In order to cut off the immune response due to Propionibacterium acnes, we took a small protein sequence of the binding site of TLR2 and P. acnes surface acylated lipoprotein for site-directed mutation to achieve the irreversible binding effect of TLR2 antagonist to acylated lipoprotein, thereby preventing the combination between acylated lipoprotein and TLR2. On the one hand, we increased the hydrophobicity of the hydrophobic pocket bound by the acylated lipoprotein, and on the other hand, we contracted the slack &#946;-loop sheet. In this way, we not only improve the hydrophobic interaction, but also prevent the bound antagonist from going off-target with the acylated lipoprotein.  
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===Usage and Biology===
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<p>In order to cut off the immune response due to <i>Propionibacterium acnes</i>, we took a small protein sequence of the binding site of TLR2 and <i>P. acnes</i> surface acylated lipoprotein for site-directed mutation to achieve the irreversible binding effect of TLR2 antagonist to acylated lipoprotein, thereby preventing the combination between acylated lipoprotein and TLR2. On the one hand, we increased the hydrophobicity of the hydrophobic pocket bound by the acylated lipoprotein, and on the other hand, we contracted the slack &#946;-loop sheet. In this way, we not only improve the hydrophobic interaction, but also prevent the bound antagonist from going off-target with the acylated lipoprotein. </p>
  
  
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===Usage and Biology===
 
  
 
===RESULTS===
 
===RESULTS===
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<center><img src="https://2021.igem.org/wiki/images/c/c7/T--HUST2-China--Heat_released_in_PBS_titration_with_100_times_diluted_formic_acid.png" style="width:60%"></center>
 
<center><img src="https://2021.igem.org/wiki/images/c/c7/T--HUST2-China--Heat_released_in_PBS_titration_with_100_times_diluted_formic_acid.png" style="width:60%"></center>
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<center><b>Figure 2.</b>Heat released in PBS titration with 100 times diluted formic acid. The upper Figure shows the heat released per second at a given point in the titration process. The lower Figure shows the heat released per drop of formic acid diluted 100 times with PBS. Add only 20μL formic acid diluted 100 times with PBS for the first drop and 40μL for the next 19 drops. At equilibrium, the titration heat released is about 1.50μcal per second and 120kcal per mole of the injected substance.</center>
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<center><b>Figure 1.</b>Heat released in PBS titration with 100 times diluted formic acid. The upper Figure shows the heat released per second at a given point in the titration process. The lower Figure shows the heat released per drop of formic acid diluted 100 times with PBS. Add only 20μL formic acid diluted 100 times with PBS for the first drop and 40μL for the next 19 drops. At equilibrium, the titration heat released is about 1.50μcal per second and 120kcal per mole of the injected substance.</center>
 
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Latest revision as of 19:57, 21 October 2021


Optimized TLR2 antagonists

Usage and Biology

In order to cut off the immune response due to Propionibacterium acnes, we took a small protein sequence of the binding site of TLR2 and P. acnes surface acylated lipoprotein for site-directed mutation to achieve the irreversible binding effect of TLR2 antagonist to acylated lipoprotein, thereby preventing the combination between acylated lipoprotein and TLR2. On the one hand, we increased the hydrophobicity of the hydrophobic pocket bound by the acylated lipoprotein, and on the other hand, we contracted the slack β-loop sheet. In this way, we not only improve the hydrophobic interaction, but also prevent the bound antagonist from going off-target with the acylated lipoprotein.


RESULTS

We find that when formic acid is diluted 100 times with PBS and titrated with PBS, the heat released is minimal and relatively stable. Under this background, the concentration of acylated lipoprotein is 60μM, and the concentration of Wild-type/Optimized TLR2 antagonist is 6μM. Therefore, we are confident of the experimental results.

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Figure 1.Heat released in PBS titration with 100 times diluted formic acid. The upper Figure shows the heat released per second at a given point in the titration process. The lower Figure shows the heat released per drop of formic acid diluted 100 times with PBS. Add only 20μL formic acid diluted 100 times with PBS for the first drop and 40μL for the next 19 drops. At equilibrium, the titration heat released is about 1.50μcal per second and 120kcal per mole of the injected substance.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 172