Part:BBa_K2910002
IsPETase-ML Mut 1 with C-terminal Hexahistidine Tag
BBa_K2910002 encodes a variant of the Ideonella sakaiensis PETase, optimized for E. coli, which degrades poly(ethylene terephthalate. Using a Machine Learning Algorithm to optimize hydrophobicity content, the thermostability of the protein is predicted to be improved.
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]
In order to assess the enzymatic activity of PETase variants relative to the wild type, and W159H S238F PETase (BBa_k2910000), p-nitrophenyl butyrate assays were performed. Ester bonds in PET plastic are mimicked by p-nitrophenol. Each PETase variant cleaves the ester bond; once the bond is broken it emits a wavelength at 405 nm. This emission can be read by a plate reader; if PETase is catalytically active, absorbance at 405 should be perceived. The following figures show the results of the aforementioned assay for this variant.
A plot of the pNB Hydrolysis at pH = 8 by BBa_K2910002 is presented below: <img src="https://parts.igem.org/File:T--Toronto--pNPB-Hydrolysis-pH-8_Tianyu-All-Conc.png">
A plot of the pNB Hydrolysis at pH = 9 by BBa_K2910002 is presented below: <img src="https://parts.igem.org/File:T--Toronto--pNPB-Hydrolysis-pH-9_Tianyu-All-Conc.png">
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