Part:BBa_K2933015
subclass B1 metallo-beta-lactamase TMB-2, codon optimized in E. coli
This part encodes a protein called TMB-2, which is a metallo-beta-lactamase of subclass B1.
The Tripoli metallo-beta-lactamase-1 (TMB-1) gene was first discovered in a Achromobacter xylosoxidans strain obtained from an environmental sample in a hospital in Tripoli, Libya, in 2011 After the initial report, TMB-1 has been identified in clinical isolates of Acinetobacter spp. in Japan, and the new TMB-1 variant named TMB-2, with the single mutation S228P, was isolated from a different hospital in Japan also in clinical isolates of Acinetobacter spp.
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]
Usage and Biology
TMB-2 is a type of subclass B metal beta-lactamases, which is derived from TMB-1 mutation. The beta lactamases of the TMB family 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 and pET-28b_SUMO to construct our expression plasmid. And then we converted the plasmid constructed to E. coli DH5α to expand the plasmid largely.
Figure 1. Left: The PCR result of TMB-2. Right: The verification results by 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.
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