Part:BBa_K5040002

ZFa-hQDPR

The fusion protein of hQDPR and ZFa binding domain. hQDPR reduces the qBH2 to the BH4, acting as an enzyme in the BH4 regeneration system.

Experimental result

After confirming the cloning, we induced the ZFa-hQDPR fusion protein expression by treating 0.02 mM IPTG at 37°C.

Fig. The Coomassie Blue staining of SDS-PAGE analysis of the ZFa-hQDPR fusion protein at 0, 4, and 8 hours of IPTG induction. Sup: supernatant; P: pellet

We aim to determine whether the protein expression level increases with extended time; therefore, we conducted inductions for 0, 12, and 24 hours.

Fig. The Coomassie Blue staining of SDS-PAGE analysis of the ZFa-hQDPR and 24 hours of IPTG induction. Sup: supernatant; P: pellet

We also want to determine whether the different IPTG concentration influence protein expression level; therefore, we conducted inductions with IPTG concentrations of 0.2 mM, 0.4mM, 0.8mM.

Fig. The Coomassie Blue staining of SDS-PAGE analysis of the ZFa-hQDPR fusion protein induced with IPTG concentrations of 0.2 mM, 0.4 mM, and 0.8 mM at 8 hours.

To achieve our goal, we builded a composite part (Part:BBa_K5040008) for 5-HTP production.

Fig. The Coomassie Blue staining of SDS-PAGE analysis of co-expressing Zif268-rlinker-m-hTPH1, PBSII-rlinker-hPCBD1, and ZFa-hQDPR fusion protein at 0, 4, and 8 hours of IPTG induction. Sup: supernatant; P: pellet

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 526
  
• 1000
  COMPATIBLE WITH RFC[1000]