Part:BBa_M36071
K12 e.coli native Manganese Superoxide Dismutase
Coding sequence for the enzyme Mn-SOD or SoDA. This protein catalyzes the dismutation of superoxide, a free radical. Sequence found by converting the amino acid sequence of Mn-SOD protein native to Escherichia coli, the organism for which it is also optimized for.
Improvement Made by Xiamen_city 2020
Our composite part BBa_K3523005 contains the basic part BBa_K3523000, which sequence has optimized differently from the sequence of other two similar basic part BBa_M36071 and BBa_K2638106 (Fig1). Apart from that, we add a T7 promoter, His tag, and a T7 terminator to develop our composite part BBa_K3523005.
There is a mini-review of the development of SOD protein related parts. In 2012, group Stanford BIOE44-S11 designed a basic part BBa_M36071, aimed to catalyze the dismutation of superoxide. Although they attempted to utilize and overexpress this protein, they did not get their expected results. In 2018, group iGEM18_Bielefeld-CeBiTec designed a basic part BBa_K2638106, which has optimized the coding sequence of SOD protein. They also designed some related composite parts, like BBa_K2638117, BBa_K2638118, and BBa_K2638116. However, all these parts did not attach any experimental data to prove the function of SOD protein. Today, our team further improved the coding sequence of SOD protein and constructed composite part BBa_K3523005. In order to prove the function of these parts, we expressed and purified SOD protein, and then detected the enzyme activity in vitro. As the result shown, our SOD protein has achieved engineering success. Besides, our project aimed to degrade reactive oxygen species (ROS) accumulated when people staying up late. And the SOD can excellent degrade ROS into H2O2 that is accord with our initial expectation.
Usage and Biology
Superoxide dismutase cataylyzes the dismutation of superoxide. Superoxide is produced naturally in the respiration process. It causes damage to the cell by altering lipid structure and breaking down proteins among other negative effects. This gene provides the protein to transform two superoxide molecules into hydrogen peroxide and oxygen.
Design Notes
This DNA sequence is converted from the amino acid sequence of Mn-SOD protein using the online converter from In-Silico. We optimized our sequence in DNA 2.0's Gene Designer by increasing GC content and minimizing repeats. We accomplished this by altering basepairs without altering amino acids.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 346
Illegal PstI site found at 283
Illegal PstI site found at 593 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 346
Illegal PstI site found at 283
Illegal PstI site found at 593 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 346
Illegal XhoI site found at 43
Illegal XhoI site found at 127 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 346
Illegal PstI site found at 283
Illegal PstI site found at 593 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 346
Illegal PstI site found at 283
Illegal PstI site found at 593
Illegal NgoMIV site found at 542 - 1000COMPATIBLE WITH RFC[1000]
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