Affinity tags on fusion proteins are widely used in protein expression techniques. The most common affinity tags, such as glutathione S-transferase (GST), polyhistidine, maltose-binding protein (MBP), and streptavidin tags, are commonly used to increase expression, improve solubility, and facilitate protein purification. High-affinity tags (MBPs, GSTs) are known to influence the conformational homogeneity and thus the three-dimensional structure of folded proteins in vivo.
The current study described the in vivo effects of the small-affinity N-terminal 6xHis-tagged Recombinant fusion on cell growth. The hexahistidine-tagged full-length β-clamp and the hexahistidine-untagged β-clamp were overexpressed and coexpressed in possible combinations with truncated DnaE in the expressing strain of E. coli. After IPTG induction, protein expression was assessed by SDS PAGE.
Comparative analysis of the growth curves generated for induced and uninduced cells demonstrated decreased growth rates of cells overexpressing the non-6xHis-tagged β-clamp compared to the 6xHis-tagged β-clamp. Based on analysis of soluble and insoluble protein fractions by SDS PAGE gels and the published His-tagged β-clamp structure (PDB: 4K74), we propose that the N-terminal 6 x His tag on the β-clamp occludes its Gly66 to ultimately affect its ability to interact with clamp loader subunit 8.
Keywords: 6xHis tag; DNA polymerase; affinity tag; clamp charger; β-clamp.
Writes: recombinant protein
STAT3 | Acute phase response factor | APRF | DNA-binding protein APRF | HIES
Label: 6His, N-terminal
Region: yy 1-770
Expression system: Yeast
Species of origin: Yeast
Purification: More than 90% by SDS-PAGE
Bio-Activity: Not tested
Endotoxin: Not tested
Presentation: 20 mM Tris-HCl, 0.5 M NaCl (pH 8.0) with 10% glycerol.
Short-term: Store at 2°C to 8°C for up to 1 week. Long Term: Aliquot and store at -20°C to -80°C. Avoid freeze-thaw cycles.
For research use only. Intended for use by laboratory professionals.
This protein carries the 100% LSBio Guarantee.
Uniprot: P40763 NCBI: NM_003150 NP_003141.2