Part:BBa_K5040001
PBSII-rlinker-hPCBD1
The fusion protein of hPCBD1 and PBSII binding domain with 36 bp rigid linker. hPCBD1 dehydrates the BH3OH to the qBH2, acting as an enzyme in the BH4 regeneration system.
Experimental result
After confirming the cloning, we induced the PBSII-rlinker-hPCBD1 fusion protein expression by treating 0.02 mM IPTG at 37°C.
Fig. The Coomassie Blue staining of SDS-PAGE analysis of the PBSII-rlinker-hPCBD1 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 PBSII-rlinker-hPCBD1 fusion protein at 0, 12, 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 Zif268-rlinker-m-hTPH1 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. After confirming the expression cassette insertion into pST39 vector, we induced the fusion proteins expression by adding 0.2mM IPTG at 37°C.
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
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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