Difference between revisions of "Part:BBa K3088001"

Latest revision as of 15:25, 21 October 2019

HBEGF/ENVZ Chimeric Complete

Heparin-binding EGF-like growth factor (HB-EGF) is a unique receptor of the diphtheria toxin. EnvZ, on the other hand, is a member of two-component regulatory system that is important for bacteria osmoregulation. When diphteria toxin binds to HB-EGF, it will phosphorylate EnvZ and activates the signalling cascade pathway. Once EnvZ is phosphorylated, OmpR (a transcription factor of OmpC promoter) will also be phosphorylated. The phosphorylated OmpR will initiate OmpC transcription and express the gene following OmpC promoter. This is the complete part containing the appropriate promoter T5 lac promoter (BBa_K592008), Ribosomal Binding Site (BBa_B0034), our HBEGF/EnvZ chimeric protein (BBa_K3088001), Histidine tag (BBa_K1223006), and double terminator (BBa_B0010 and BBa_B0012)

Usage and Biology

This part when combined with OMPC/GFP part (BBa_K769001) can be used for diphteria toxin detection with observable green fluorescent protein.

Consideration of using TOP10 bacteria in LB medium for characterizing HBEGF/EnvZ - OmpC/GFP

There are several considerations why we use TOP10 bacteria in LB medium for HE-OG characterization with heparin. Firstly, our modelling was designed based on K12 bacteria which includes TOP10. We did not include DH5a because the results of the OMPC-GFP (BBa_K769001) expression in DH5a showed inconsistencies when the sodium concentration is increased. TB medium and low salt medium was also omitted to exclude any conditions which may affect the basal metabolism of the bacteria because these mediums are modified medium less suitable for optimal bacteria environment. Therefore, in order to measure the HE-OG expression for the very first time, we would like to use optimal environment (LB medium) as well as TOP10 bacteria to maximize the result.

Sequence and Features