Part:BBa_K4117888

UGT1A3

“T7 promoter+ pmrA+ RBS+ pmrB+ rrnB T1 terminator+ pmrC promoter” is the THC sensing pathway. To alert vaccinators that they have ingested Δ9-THC, we design a Δ9-THC sensing system. In this pathway, anti-THC can specifically bind to Δ9-THC, and then promote PmrB phosphorylation of PmrA. PmrC promoters activate phosphorylated PmrA proteins and initiate transcription to express UGT1A3.

Outer membrane protease T (ompT) is a 33.5 kDa endoprotease located on the outer membrane of Escherichia coli. OmpT signal peptide is an excretion tag linked on the N-terminus of protein. It can transport proteins across the outer membrane with the help of ompT.

UGT1A3 in this pathway plays a significant role in metabolic section, working upon the sensing section. Tetrahydrocannabinol (Δ9 -THC), the primary psychoactive ingredient in marijuana, is subject to cytochrome P450 oxidation and subsequent UDP-glucuronosyltransferase(UGT)-dependent glucuronidation. Many studies have shown that CYP2C9 and CYP2C19 are the primary enzymes responsible for these cytochrome P450-dependent oxidations. In Mazur’s study, the specific human UGTs are responsible for classic cannabinoid metabolism, and among them, the highest activity toward THC-COOH was observed with UGT1A3.

Thus, in our system, Δ-9-THC is hydroxylated by CYP2C9(BBa_K4117002) and CYP2C19(BBa_K4117022), remaining psychoactive. The major product of CYP2C9 and CYP2C19 metabolism, 11-OH-Δ-9-THC, was further oxidized to THC-COOH. Then UGT1A3, which catalyzes the glucuronidation of the former substrate, increases the metabolite's water solubility, thereby facilitating excretion into the urine.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 4599
  Illegal XbaI site found at 693
  Illegal SpeI site found at 713
  Illegal PstI site found at 1214
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 4599
  Illegal SpeI site found at 713
  Illegal PstI site found at 1214
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 4599
  Illegal BglII site found at 4257
  Illegal BamHI site found at 2887
  Illegal BamHI site found at 3444
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 4599
  Illegal XbaI site found at 693
  Illegal SpeI site found at 713
  Illegal PstI site found at 1214
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 4599
  Illegal XbaI site found at 693
  Illegal SpeI site found at 713
  Illegal PstI site found at 1214
  Illegal AgeI site found at 3031
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 732