Difference between revisions of "Part:BBa K4165010"
 
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<partinfo>BBa_K4165010 short</partinfo>
 
<partinfo>BBa_K4165010 short</partinfo>
  
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This basic part encodes Human serine protease inhibitor known as SPINK8 which is able to inhibit trypsin-like proteases, like HtrA1 (BBa_K4165004).
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This basic part encodes Human serine protease inhibitor known as SPINK8 which is able to inhibit trypsin-like proteases like serine protease HtrA1 (BBa_K4165004).
  
 
===Usage and Biology===
 
===Usage and Biology===
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This type of family encodes for a type of inhibitor that
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This type of family encodes inhibitors that is predicted to be able to inhibit serine peptidases. The inhibitor is present extracellularly and binds to trypsin-like proteases (serine proteases) and since the catalytic core of HtrA1 (BBa_K4165004) is considered as a trypsin-like catalytic domain, so this inhibitor also is considered to inhibit the function of HtrA1.
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is predicted to be able to inhibit serine peptidases. The inhibitor is present  
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extracellularly. The inhibitor binds to trypsin-like proteases (serine proteases)  
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and since the catalytic core of HtrA1 (BBa_K4165004) is considered as a trypsin-like  
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catalytic domain, so this inhibitor also is considered to inhibit the  
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function of HtrA1 <sup>[1-3]</sup>.
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===<span class='h3bb'>Sequence and Features</span>===
 
===<span class='h3bb'>Sequence and Features</span>===
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===Functional Parameters===
 
===Functional Parameters===
  
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GC% Content
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<html>
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61.2%
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<style>
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table, th, td {
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  border:1px solid black; margin-left:auto;margin-right:auto;
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}
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</style>
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<body>
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<table style="width:65%">
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<table>
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  <tr>
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    <th>GC Content%</th>
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    <th>Isoelectric point (PI)</th>
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    <th>Charge at pH 7</th>
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    <th>Molecular Weight (Protein)</th>
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  </tr>
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  <tr>
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    <td>61.2%</td>
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    <td>4.597</td>
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    <td>-3.459</td>
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    <td>10.821</td>
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  </tr>
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</table>
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</body>
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</html>
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===Dry Lab Characterization===
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<p style=" font-weight: bold; font-size:14px;"> Modeling </p>
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This inhibitor was modeled using different software (trRosetta - AlphaFold2 - RosettaFold - Modeller) and the top model was acquired from trRosetta ranking 6 out of 6 according to our Quality Assessment code.
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<html>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/p0c7l1-trrosetta-model2.png" style="margin-left:200px;" alt="" width="500" /></p>
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</html>
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                            Figure 1. Predicted 3D structure of SPINK8 inhibitor Visualized on Pymol.
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<p style=" font-weight: bold; font-size:14px;"> Docking </p>
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ΔG = -25.0
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<html>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/parts-registry/htra1/htra1-p0c7l1.jpeg" style="margin-left:200px;" alt="" width="500" /></p>
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</html>
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                          Figure 2. Docked structure of HtrA1 with SPINK8 inhibitor Visualized on Pymol.
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<p style=" font-weight: bold; font-size:14px;"> Mathematical modeling </p>
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<p style=" font-weight: bold; font-size:14px;">Transcription rate and translation rate under T7 promotor </p>
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the mathematical modeling was based on our code for the calculation of transcription and translation (you can find it in the code section) beside with the estimated results from the wet lab.
  
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Isoelectric point (PI)
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<html>
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4.597
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<p><img src="https://static.igem.wiki/teams/4165/wiki/dry-lab/mathematical-modeling/mathematical-modeling/p0c7l12.png" style="margin-left:200px;" alt="" width="500" /></p>
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</html>
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Charge at pH 7
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                Figure 3. this figure shows the results from the transcription and translation code showing the
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-3.459
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                    variation of mRNA and protein concentrations with time compared with the wet lab results.
  
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Molecular Weight (Protein)
 
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10.821 kDa
 
  
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===PDB Structure===
 
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It is a predicted model (AlphaFold2).
 
  
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AlphaFold:
 
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https://alphafold.ebi.ac.uk/entry/P0C7L1
 
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Molprobity:
 
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Clash Score:
 
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Ramachandran Favoured:
 
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Ramachandran Outliers:
 
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Rotamers Outliers:
 
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C-beta Deviations:
 
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Q-Mean:
 
  
 
===References===
 
===References===

Latest revision as of 13:54, 13 October 2022


Human serine protease inhibitor Kazal type 8 (SPINK8)

This basic part encodes Human serine protease inhibitor known as SPINK8 which is able to inhibit trypsin-like proteases like serine protease HtrA1 (BBa_K4165004).

Usage and Biology

This type of family encodes inhibitors that is predicted to be able to inhibit serine peptidases. The inhibitor is present extracellularly and binds to trypsin-like proteases (serine proteases) and since the catalytic core of HtrA1 (BBa_K4165004) is considered as a trypsin-like catalytic domain, so this inhibitor also is considered to inhibit the function of HtrA1.

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
    COMPATIBLE WITH RFC[1000]

Functional Parameters

GC Content% Isoelectric point (PI) Charge at pH 7 Molecular Weight (Protein)
61.2% 4.597 -3.459 10.821


Dry Lab Characterization

Modeling

This inhibitor was modeled using different software (trRosetta - AlphaFold2 - RosettaFold - Modeller) and the top model was acquired from trRosetta ranking 6 out of 6 according to our Quality Assessment code. 



                            Figure 1. Predicted 3D structure of SPINK8 inhibitor Visualized on Pymol.


Docking

ΔG = -25.0 

                          Figure 2. Docked structure of HtrA1 with SPINK8 inhibitor Visualized on Pymol.

Mathematical modeling

Transcription rate and translation rate under T7 promotor

the mathematical modeling was based on our code for the calculation of transcription and translation (you can find it in the code section) beside with the estimated results from the wet lab. 


               Figure 3. this figure shows the results from the transcription and translation code showing the 
                    variation of mRNA and protein concentrations with time compared with the wet lab results.



References

1 - Frochaux, V., Hildebrand, D., Talke, A., Linscheid, M. W., & Schlüter, H. (2014). Alpha-1-antitrypsin: a novel human high temperature requirement protease A1 (HTRA1) substrate in human placental tissue. PloS one, 9(10), e109483. 2 - Grau, S., Baldi, A., Bussani, R., Tian, X., Stefanescu, R., Przybylski, M., ... & Ehrmann, M. (2005). Implications of the serine protease HtrA1 in amyloid precursor protein processing. Proceedings of the National Academy of Sciences, 102(17), 6021-6026. 3 - Eigenbrot, C., Ultsch, M., Lipari, M. T., Moran, P., Lin, S. J., Ganesan, R., ... & Kirchhofer, D. (2012). Structural and functional analysis of HtrA1 and its subdomains. Structure, 20(6), 1040-1050.