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Thermo Scientific WELQut Protease is highly specific, recombinant serine protease of Staphylococcus aureus. It recognizes and precisely cleaves recombinant proteins containing an engineered recognition sequence† W- E- L- Q↓X (Trp, Glu, Leu, Gln, X can be any amino acid). The protease cleaves outside the recognition sequence without leaving additional amino acids bound to the target protein.
The WELQut Protease is active in a broad temperature (4 to 30°C) and pH (pH 6.5 to 9.0) range and does not require specific buffers.
In addition, this new protease has several procedural advantages - it is ideal for on-column proteolysis reactions and can be easily removed from reaction mixtures using its built-in His-tag.
† This cleavage sequence is present in expression vector pLATE52 included into the Thermo Scientific aLICator LIC Cloning and Expression Kit 4 (N-terminal His-tag/WELQut) (#K1281) available from Thermo Scientific.
Figure 1 | Highly specific proteolysis of target protein performed by WELQut vs. competitor Enterokinase. Target protein (Klenow Fragment (exo-), containing WELQut or Enterokinase recognition sequence) was treated with WELQut Protease and Enterokinase according to the recommendations provided by the suppliers. Various enzyme/substrate amount ratios were tested and reaction products were analyzed in SDS-PAGE.M - PageRuler Prestained Protein Ladder (Thermo Scientific, #SM0671).K - Control-uncut target protein.1-6 various WELQut/target protein amount ratios, U/µg.1 - 1:100, 2 - 1:50, 3 - 1:40, 4 - 1:20, 5 - 1:10, 6 - 1:5. 7-11 various Enterokinase/target protein amount ratios, µg/µg.7 - 1:100, 8 - 1:40, 9 - 1:20, 10 - 1:10, 11 - 1:5.
Figure 2 | Ideal for on-column proteolysis reactions. Target protein (Klenow Fragment (exo-)), containing His-tag and WELQut recognition sequence, was treated with WELQut on-column, during IMAC purification procedure. (HisPur Ni-NTA Spin Columns, Thermo Scientific Pierce).M - PageRuler Prestained Protein Ladder (Thermo Fisher Scientific, #SM0671).1 - negative control (E.coli ER2566/pLATE52-Cat before induction).2 - lysate load (E.coli ER2566/pLATE52-Klenow Fragment after 3 h induction with 0.1 mM IPTG.3 – Eluate I (lane shows that after enzymatic His6 tag removal, Klenow Fragment is eluted from the IMAC sorbent).4 – Eluate II - imidazole eluted proteins (lane shows cleavage reaction and removal of WELQut from reaction mixture efficiencies).