Coding

Part:BBa_K4165051

Designed by: Esraa Elmligy   Group: iGEM22_CU_Egypt   (2022-09-30)
Revision as of 14:29, 13 October 2022 by M Zaki (Talk | contribs) (Dry-Lab characterization)


HtrA1 Switch 31

This composite part consists of T7 promoter (BBa_K3633015), lac operator (BBa_K4165062), pGS-21a RBS (BBa_K4165016), 6x His-tag (BBa_K4165020), H1A (BBa_K4165000), GS Linker (BBa_K4165066), seed peptide (BBa_K4165012), GS Linker (BBa_K4165019), seed peptide (BBa_K4165012), GS Linker (BBa_K4165066), WAP inhibitor (BBa_K4165008), and T7 terminator (BBa_K731721).

Usage and Biology

Switch 31 is used to mediate the activity of HTRA1. It is composed of 3 parts connected by different linkers; an HtrA1 PDZ peptide, a clamp of two targeting peptides for tau or amyloid beta, and a catalytic domain inhibitor. Activating HTRA1 requires a conformational change in the linker, eliminating the attached inhibitor from the active site. The conformational rearrangement can be mediated through the binding of affinity clamp to tau or beta-amyloid. This binding will result in a tension that detaches the inhibitor from the active site.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 589
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 572
    Illegal AgeI site found at 308
  • 1000
    COMPATIBLE WITH RFC[1000]

Dry-Lab characterization

Modeling

The switch was modeled by (Alphafold - Rosettafold - tRrosetta) and the top model was obtained from tRrosseta.

cbeta_deviations clashscore molprobity ramachandran_favored ramachandran_outliers Qmean_4 Qmean_6
2 1.55 89.84 1.56 0.195965 0.384058 0.196775

                            Figure 1. The 3D structure of switch 31 modeled by TRrosetta
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Categories
Parameters
None