Part:BBa_K1151000
Nickel-responsive pleiotropic regulator (HpNikR)
The HpNikR protein is a pleiotropic regulator from Helicobacter pylori. In presence of nickel it can acts as an activator or a repressor depending of the specific promoter that contains its operator site. It consists of two dimeric DNA binding domains separated by a tetrameric regulatory domain that binds nickel. This domain corresponds to the C-terminal regulatory domain which contains four nickel binding sites at the tetramer interface. Binding nickel, then a conformational change allows it to activate or repress trascription.
Usage and Biology
Figure 1: HpNikR nickel-binding domain (tetramer).
Figure 2: Proteic sequence analysis with Uniprot.
NikR expression using BL21 (DE3) cells
First we made competent BL21 cells and we transformed it with the plasmid containing NikR. We then proceeded with the normal protocol of induction with IPTG.
Figure 3: PAGE of the first induction.
Cytosol/membrane separation by Zerial method
To confirm that NikR is a cytosolic protein (not expressed in multivesicular bodies, and then in membrane) we performed a separation membrane-cytosol (Zerial method) (sample: 2-hours induced cells).
Figure 4: PAGE of the separation.
NikR purification by Ni-NTA resin
His-tagged NikR can be purified by Ni-NTA resin, which has a high affinity for histidine residues.
Figure 5: Purification result
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Unknown
- 23INCOMPATIBLE WITH RFC[23]Unknown
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
| n/a | Nickel-responsive pleiotropic regulator (HpNikR) |