Coding

Part:BBa_K4949004

Designed by: UManitoba 2023   Group: iGEM23_UManitoba   (2023-10-10)
Revision as of 03:04, 12 October 2023 by Prefont5 (Talk | contribs)

Est119

Est119 is an esterase originally identified in the Thermobifida alba strain AHK119 (AB298783). Est119 has been shown to degrade aliphatic-aromatic copolyesters and decrease the size of polymer particles of other biodegradable plastics, with an optimal temperature range of 45-55°C (Hu and al. 2009). Similarly to MGS0156, Est119 is interesting due to its potential temperature compatibility with Manitoban composting methods. Our team modified the Est119 genetic sequence to include the Lpp-OmpA anchor to allow for the characterization of the surface-display mechanism of PLAnet Zero

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]



Figure 1. Absorbance as a function of time for the whole cell esterase activity assay of Est119. After induced expression, E.coli BL21 (DE3) + Est 119 or E.coli BL21 (DE3)ΔLpp + Est119, a native Lpp deficient stain, were washed and resuspended in PBS with 100µM NPO with a cell density normalized to OD600=1.0. Samples were then incubated for 5 min at room temperature. Aliquots of 100 µl were collected at 5 intervals and the absorbance of the supernatant as measured at 410 nm.

Figure 2. Absorbance as a function of time for the whole cell esterase activity assay of Est119. After induced expression, E.coli BL21 (DE3) + Est119 or E.coli BL21 (DE3)ΔLpp +Est119, a native Lpp deficient stain, were washed and resuspended in PBS with 100µM pNOB with a cell density normalized to OD600=1.0. Samples were then incubated for 5 min at room temperature. Aliquots of 100 µl were collected at 5 intervals and the absorbance of the supernatant as measured at 410 nm.

Figure 3. Absorbance as a function of time for the whole cell esterase activity assay of Est119. After induced expression E.coli BL21 (DE3) + Est119 or E.coli BL21 (DE3) + Est119(ΔLpp), bacteria with a Lpp deficient plasmid, were washed, and resuspended in PBS with 100µM pNOB with a cell density normalized to OD600=1.0. Samples were then incubated for 5 min at room temperature. Aliquots of 100 µl were collected at 5 intervals and the absorbance of the supernatant as measured at 410 nm.

Figure 4. Absorbance as a function of time for the whole cell esterase activity assay of Est119. After induced expression, E.coli BL21 (DE3) + Est119 or E.coli BL21 (DE3) + Est119(ΔLpp), bacteria with an Lpp deficient plasmid, were washed and resuspended in PBS with 100µM NPO with a cell density normalized to OD600=1.0. Samples were then incubated for 5 min at room temperature. Aliquots of 100 µl were collected at 5 intervals and the absorbance of the supernatant as measured at 410 nm.

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