Part:BBa_K2933021
subclass B1 metallo-beta-lactamase Fla.103, codon optimized in E. coli
This part encodes a protein called Fla.103, which is a metallo-beta-lactamase of subclass B1.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 76
- 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 76
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 558
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 76
- 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 76
- 1000COMPATIBLE WITH RFC[1000]
Usage and Biology
Fla.103 is a type of subclass B metal beta-lactamases. The beta lactamases can hydrolyze almost all available beta lactam antibiotics (except aztreonam) clinically, including the broad-spectrum antibiotic carbapenems. Because of the extensive substrate profile of this enzyme, the clinical strains carrying it become a great threat to human life and health.
References
Molecular cloning
First, we used the vector pGEX-6p-1 to construct our expression plasmid. And then we converted the plasmid constructed to E. coli DH5α to expand the plasmid largely.We then extracted plasmids and performed double enzyme digestion verification.
Figure 1. (a) The PCR result of Fla.103. (b) The verification results by enzyme digestion.The 1 is the original plasmid.The 2 is the
results verified by double enzyme digestion.
After verification, it was determined that the construction is successful. We converted the plasmid to E. coli BL21(DE3) for expression and purification.But we failed.
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