Difference between revisions of "Part:BBa K4165030"

Revision as of 11:48, 10 October 2022

HtrA1 switch 10

This composite part consists of T7 promoter (BBa_K3633015), lac operator (BBa_K4165062), pGS-21a RBS (BBa_K4165016), 6x His-tag (BBa_K4165020), H1A peptide (BBa_K4165000), GGGGSG (BBa_K4165017), TD28rev (BBa_K4165006), GGSGGGG (BBa_K4165018), WWW (BBa_K4165007), SPINK8 inhibitor (BBa_K4165010) and T7 terminator (BBa_K731721).

Source

Synthesized

Usage and Biology

Switch 10 is used to mediate the activity of HTRA1. 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 affinity clamp for tau and beta-amyloid binding. These clamps are used for stabilizing the inhibitor away from the active site. These two domains (inhibitor and affinity clamp connected with linker1). Additionally, (H1A) binding peptide bound to the PDZ domain and connected to the affinity clamp on the other side with linker3.

Dry Lab

Modeling

TRrosetta models this composite part with a score of 5 out of 6 according to our quality assessment code (you can find the python script file on the programming club page with further explanation of how you can optimize it to your needs).

                              Figure 1. The 3D structure of switch 10 modeled by TRrosetta. Red: Tau binding peptides, 
                                              blue: H1A peptide, cyan: inhibitor and green: linkers

Docking

switch 10 vs HtrA1 trimer:

ΔG = -20.041

                              Figure 2. The 3D structure of switch 10 docked to HtrA1

Mathematical modeling

Transcription rate and translation rate

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.