Expression and purification of protein using Strep-Tag and /or 6xHistidine-tag

Expression and purification of protein using Strep-Tag and /or 6xHistidine-tag

Download Free online Expression and purification of protein using Strep-Tag and /or 6xHistidine-tag Manual PDF

The Strep-tag® II is a short peptide (8 amino acids, WSHPQFEK), which binds with high selectivity to Strep-Tactin®, an engineered streptavidin. The binding affinity of Strep-tag® II to Strep-Tactin® (Kd= 1 μM) is nearly 100 times higher than to streptavidin. This technology allows one-step purification of almost any recombinant protein under physiological conditions, thus preserving its bioactivity. The Strep-tag® system can be used to purify functional Strep-tag® II proteins from any expression system including baculovirus, mammalian cells, yeast, and bacteria [1,2,3].Twin-Strep-tag®(former One-STrEP-tag)Based on the proprietary Strep-tag® technology the Twin-Strep-tag®was developedwhich has a size of 3kDa.TheTwin-Strep-tag® combines high specificity and mild conditions with higher affinity thereby enabling efficient purification even in batch or directly from culture supernatants. Furthermore, it tolerates diverse buffer conditions and additives (high salt, detergents, reducing agents, metal ions and chelating agents) making it a universal tag for varying protein properties, particularly for protein complexes in protein interaction analysis.After application of the crude extract on a Strep-Tactin® column and a short washing step, gentle elution of purified recombinant protein is performed by addition of low concentrations (2.5 mM) desthiobiotin. The Strep-tag®/Strep-Tactin® interaction is compatible with a variety of reagents (seeTable 1) making the system attractive for purifing metallo-and membrane proteins, large proteins and protein complexes. Binding capacity depends on the Strep-Tactin® matrices (25 -100 nmol/ml using standard Strep-Tactin® resins and up to 500nmol/ml using Strep-Tactin® Superflow® High Capacity) as well ason the fused recombinant protein.Because of its small size, Strep-tag® generally does not interfere with the bioactivity of the fusion partner. Thus, removal of the tag becomes superfluous. Comprehensive reviews and scientific publications giving an overview of various Strep-tag®applications are listed at resin?If you are unsure which resin is the right choice for your protein, test our different Strep-Tactin®resins to evaluate thebest one for the purification of your protein of interest. For rapid evaluation we are offering two Column Evaluation Setswith Strep-Tactin®immobilized to the supports Sepharose, Superflow and MacroPrep,respectively. The Sets areoffered either with 0.2 ml Mini-columns( 2-1001-011)or 1 ml gravity flow columns ( 2-1001-001).

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