Part:BBa_K2632008
Plac-Lpp-OmpA-RGD
We gave the surface display system from a new function through displaying a RGD motif on the Lpp-OmpA which contains a signal sequence, the N-terminal of the lipoprotein (Lpp) and the residual 46-159 amino acids of the OmpA. In addition, lipoprotein executes the function of targeting to the outer membrane and OmpA constructs an anchor on the outer membrane. RGD motif can specifically bind to αVβ3, a biomarker of cancer cells such as melanoma, neuroblastoma, glioma, adenocarcinoma1. We determine the surface display site on the third loop of Lpp-OmpA through homology modelling (Figure 1). This part is under the control of lac promoter.
Figure 1. The homology modelling result of Lpp-OmpA-RGD. Red arrow shows the location of RGD motif.
Microscopy shows that Lpp-OmpA-RGD overexpressed by E. coli with 0.1mM IPTG can bind to αVβ3-positive MDA-MB-231 cell. Red arrow point the location of E. coli (Figure 2). But can not bind to αVβ3-negative MCF7 cell (Figure 3). We also use BBa_J36850 as a control. This strain can not bind to Vβ3-positive MDA-MB-231 cell line (Figure 4) and αVβ3-negative MCF7 cell line (Figure 5). These results suggest that we successfully improve the part BBa_J36850.
Figure 2. αVβ3-positive MDA-MB-231 cell line was incubated with E. coli which constructively expressed RFP and contained BBa_J36850. This improved part expressed RGD motif under the control of lac promoter.
Figure 3. αVβ3-negative MCF7 cell line was incubated with E. coli which constructive expressed RFP and contained BBa_J36850. This improved part expressed RGD motif under the control of lac promoter.
Figure 4. αVβ3-positive MDA-MB-231 cell line was incubated with E. coli which constructively expressed RFP and contained BBa_J36850.
Figure 5. αVβ3-negative MCF7 cell line was incubated with E. coli which constructively expressed RFP and contained BBa_J36850.
Reference
1. Park, S. H. et al. RGD peptide cell-surface display enhances the targeting and therapeutic efficacy of attenuated Salmonella-mediated cancer therapy. Theranostics 6, 1672–1682 (2016).
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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