Part:BBa_K3111401
TmEnc_StrepII + sfGFP_TP
T. maritima encapsulin co-expressed with sfGFP in order to measure loading capacity.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 183
Illegal BglII site found at 547
Illegal BamHI site found at 1942 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 962
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 1167
Illegal BsaI.rc site found at 2162
Illegal SapI.rc site found at 481
Illegal SapI.rc site found at 512
Experimental Results
To measure encapsulin loading, spectrophotometric analysis of purified sample was performed. The sample was excited at 280 nm and 485 nm, finding total protein concentration and fluorescent intensity of the sample. Then, a calibration curve was used to determine sfGFP concentration, adjusting for interference. Finally, this concentration was subtracted from the total protein concentration, in order to get encapsulin concentration. Concentrations were converted to molarity, and a ratio between encapsulins and cargo molecules was calculated.
From Figure 1, we can see that BBa_K3111401 loaded quite poorly, with only about 2 sfGFP molecules per encapsulin. In contrast, BBa_K3111402 loaded around 14.5 sfGFP molecules, therefore, we chose to use BBa_K3111003 over BBa_K3111002 as the building block of our delivery system due to increased loading efficiency.
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