Coding

Part:BBa_K5302005

Designed by: Dekun Zhou   Group: iGEM24_USTC   (2024-10-01)


miniZ

This year, the USTC iGEM team has utilized the competitive binding of vascular endothelial growth factor (VEGF) to develop a targeted bacterial therapy for solid tumors. Our quest for the optimal VEGF-binding protein(or peptide) led us to an in-depth exploration of proteins structurally akin to the vascular endothelial growth factor receptor (VEGFR), which we have named VEGFR-like. This part is derived from three helix 58-residue Z-domain of staphylococcal protein A. And through stabilizing mutations and the addition of a disulfide constraint the Z-domain is reengineered into a two-helix 34-residue “mini-Z” version that retains the parent's affinity. This is supposed to be more potent binders against VEGF. We used pBBR1MCS-2 plasmid as a backbone and transfered miniZ into Escherichia coli Nissle 1917, and finally succeeded in expressing miniZ.

Jamboree Program
Figure 1. sequence of miniZ and its KD with VEGF

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Figure 2. Cartoon representation of the crystal structure of the mini-Z highlighting randomized residues shown as sticks and coloring the helices

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Figure 3. Colony PCR results of pBBR-INP-miniZ

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Figure 4. SDS-PAGE analysis of pBBR1MCS-INP-miniZ expression in Escherichia coli Nissle 1917

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Figure 5. Colony PCR results of pBBR-OmpA-miniZ

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Figure 6. SDS-PAGE analysis of pBBR1MCS-OmpA-miniZ expression in Escherichia coli Nissle 1917 (1)

Jamboree Program
Figure 7. SDS-PAGE analysis of pBBR1MCS-OmpA-miniZ expression in Escherichia coli Nissle 1917 (2)


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]


[edit]
Categories
//cds/receptor
Parameters
biologyEscherichia coli Nissle 1917