Part:BBa_K4759202
T7-RBS3-OleP
P450 enzyme CYP107D1 derived from Streptomyces, abbreviated as Olep, is selected to synthesize the above gene by adding histidine tags at the C-terminus through E. coli codon optimization and subcloning the above genes onto the plasmid pET-28a to obtain the recombinant plasmid pET-28a-oleP.
Usage and Biology
OleP, the cytochrome P450 epoxidase from Streptomyces antibioticus involved in Oleandomycin biosynthesis: functional analysis and crystallographic structure in complex with clotrimazole. PDB DOI: https://doi.org/10.2210/pdb4XE3/pdb Classification: OXIDOREDUCTASE Organism(s): Streptomyces antibioticus Expression System: Escherichia coli BL21(DE3)
we selected high-copy plasmids pRSFDuet, medium-high copy plasmids pETDuet and pET28a, and low-copy plasmid pACYCDuet as carriers to express Olep in E. coli, and the results showed that the recombinant strains constructed with high-copy plasmid pRSFDuet as the carrier had the highest expression of p450 enzyme, cell growth rate, and substrate conversion rate.
Fig1: Selection of suitable plasmid. WT represents E. coli O1 strain. The blue-filled triangle represents the biomass (OD600). The red hollow triangle represents the conversion rate (%). Values and triangles represent the means and standard deviations of biological triplicates.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal XbaI site found at 47
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23INCOMPATIBLE WITH RFC[23]Illegal XbaI site found at 47
- 25INCOMPATIBLE WITH RFC[25]Illegal XbaI site found at 47
Illegal AgeI site found at 103
Illegal AgeI site found at 201 - 1000COMPATIBLE WITH RFC[1000]
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