Poly-histidine-tag

Jalili, P.R. Sharma, D. Ball, H.L. Enhancement of ionization efficiency and selective enrichment of phosphorylated peptides from complex protein mixtures using a reversible poly-histidine tag. J. Am. Soc. Mass Spectrom. 2007,18, 1007-1017. 

SR the enzyme can be purified from erythrocytes [20] or as recombinant enzyme from bacteria [9,21,22]. Preferentially, we expressed the rat SR-cDNA as N-terminally poly histidine (His6)-tagged protein in bacteria and purified it upon applying a commercially available nickel-nitrilotriacetic acid affinity column (Neuheiser, Blau and Thony, unpublished). The noncleaved, highly active fusion protein is stored in liquid nitrogen at a concentration of 1 mg/ml and an activity of approximately 1 U/ml. 

A variety of useful protein tags are available. A common one is a histidine tag, often just a string of six His residues. A poly-His sequence binds quite tightly to metals such as nickel. If a protein is cloned so that its sequence is contiguous with a His tag, it will have the extra His residues at its carboxyl terminus. The protein can then be purified by chromatography on columns with immobilized nickel. These procedures are convenient but require caution, because the additional amino acid residues in an epitope or His tag can affect protein activity. 

The absolute control over the genes that encode for the proteins readily allows the introduction of different functionalities that can be attached to synthetic polymers. Kopecek et al. [88,89] used this methodology to produce coiled-coil peptide sequences, e.g. (VSSLESK)n containing a histidine tag on the end. The hydrophobic interaction between the side groups of the valines and leucines in the sequence causes the protein to as-siune a helical or coil type structure. These hydrophobic interactions then cause several coils to further aggregate to form so called coiled coils. This coiled coil aggregation can be disrupted by temperature, pH and solvent. A copolymer of poly[N-(2-hydroxy-propyl) methacrylamide- co-(N, N"-dicarboxymethylaminopropyl)methacrylamide], (p[HPMA-co-DAMA]) with... 

Purification of polymers is performed by standard techniques that have previously been established for recombinant proteins. Frequently these include the use of affinity tags [e.g., poly(histidine), glutathione-s-transferase] that can be used to chromatographically purify recombinant proteins. If necessary, the tag can subsequently be removed by enzymatic cleavage. 

Chaga G., Bochkariov D.E., Jokhadze G.G., Hopp J. and Nelson P. Natural poly-histidine affinity tag for purification of recombinant proteins on cobalt(II)-carboxymethylaspartate crosslinked agarosa, J. Chromatogr. A, 864, 1999, 247. 

Figure 10.18 Protocol for CPNW fabrication. (Reprinted with permission from Biosensors and Bioelectronics, Label-free detection of cupric ions and histidine-tagged proteins using single poly(pyrrole)-NTA chelator conducting polymer nanotube chemiresistive sensor by C. L. Aravinda, S. Cosnier, W. Chen et ai, 24, 5. Copyright (2009) Elsevier Ltd)...

Figure 3.8 The most frequently used methods to immobilize capture molecules onto microarray surfaces. (A) Antibody adsorption by poly-L-lysine-, nitrocellulose-, or polyvinylidene fluoride-treated surface. (B) Covalent binding using various silane reagents of APTES (3-aminopropyl)triethoxysilane, GPTS (3-gly-cidoxypropyl)trimethoxysilane, and MPTS (3-mercaptopropyl)trimethoxysilane. (C) Affinity interactions by biotin/streptavidine or histidine-tag/nickel-nitrilotri-acetic acid. (D) Diffusion-based (hydrogel) antibody-immobilization technique.

Immobilized metal ion-affinily chromatography (IMAC) was first established as a technique to fractionate proteins on solid supports based on their differential affinity towards immobilized metal ions. This differential affinity derives from the eoordination bonds formed between metal ions and certain amino acid side ehains exposed on the surface of the protein molecules. Since the interaetion between the immobilized metal ions and the side chains of amino acids has a readily reversible character, it can be used for adsorption and then be disrupted under mild conditions, usually by adding a competing agent. ° The use of this technique to purify recombinant proteins containing a short affinity-tag consisting of poly-histidine residues represent its main application (the most widespread and versatile strategy used to purify recombinant proteins). ... 

Kumar A, Kamihira M, Galaev lY, lijima S, Mattiasson B (2003) Binding of Cu(II)-poly(N-isopropylacrylamide/vinylimidazole) copolymer to histidine-tagged protein a surface plasmon resonance study surface plasmon resonance study. Langmuir 19 865-871... 

Haddour N, Cosnier S, Gondran C (2005) Electrogeneration of a poly(pyrrole)-NTA chelator film for a reversible oriented immobilization of histidine-tagged proteins. J Am Chem Soc 127 5752-5753... 

While many such tag sequences exist, the most popular is a poly-6-histidine that readily binds via a chemiselective process to an immobilized metal ion chelating colnmn or IMAC (64,65). 

---