Shop All Tag Cleavage Enzymes

EKMax™ Enterokinase (Invitrogen™)

EKMax™ is a recombinant preparation of the catalytic subunit of bovine enterokinase (1). EKMax™ recognizes the sequence Asp-Asp-Asp-Asp-Lys and cleaves the peptide bond after the lysine residue. The enzyme can be used to cleave any fusion protein that carries this peptide sequence (Figure 1).

Application: Removal of fusion tags from recombinant proteins.

Unit Definition: One unit of EKMax™ is the amount of enzyme required to digest 20 µg of a thioredoxin-CAT fusion protein to 90% completion in 16 hours at 37°C. One EKMax™ unit is equivalent to ~190 trypsinogen activation units.

Pierce™ HRV 3C Protease Solution Kit (2 units/µL) (Thermo Scientific™)

Thermo Scientific Pierce Human Rhinovirus (HRV) 3C Protease is a recombinant cysteine protease used to remove fusion tags from proteins with the HRV 3C cleavage sequence and is dual tagged for easy removal from the sample after cleavage.

Features of Thermo Scientific Pierce HRV 3C Protease:

Effective—HRV 3C is designed to remove purification tags from your protein of interest
High activity at 4°C—activity is ≥2000U/mg control protein; one unit will cleave 0.1 mg control protein at 4°C after 16h
Flexible—dual-tagged enzyme is compatible with various columns and buffers

Properties:
• Alternative names: Human Rhinovirus 3C
• Source: Escherichia coli (E.coli)
• Molecular weight: 47.8kD
• Form: Liquid
• Protease type: Cysteine
• Optimal reaction conditions: 50 mM Tris-HCl, 150 mM NaCl, pH 8.0

Pierce HRV 3C Protease is highly specific for the recognition sequence Leu-Glu-Val-Leu-Phe-Gln-↓-Gly-Pro and cleaves after the glutamine residue. The recombinant enzyme is engineered as a fusion protein with glutathione-S-transferase (GST) and polyhistidine (6xHis) tags, so that the protease can be easily removed after cleavage using a variety of solid supports with immobilized glutathione (GSH) or metal chelates, respectively. HRV3C is also active in a variety of commonly used buffers, providing further flexibility in experimental design.

Applications:
• Removal of purification tags from fusion proteins

More Product Data
Choosing a vector and purification method for in vitro protein expression

AcTEV™ Protease (Invitrogen™)

AcTEV™ Protease specifically recognizes a seven amino acid sequence (Glu-Asn-Leu-Tyr-Phe-Gln-Gly, cleaving between Gln and Gly), making it useful for removing affinity tags from fusion proteins. AcTEV™ Protease is an improved version of Tobacco Etch Virus (TEV) protease that is highly site-specific, highly active, and significantly more stable than native TEV protease, resulting in enhanced long-term activity. AcTEV™ Protease features:

• Highly specific cleavage activity
• Enhanced enzyme stability for prolonged protease activity (see figure)
• Activity over a broad temperature (+4°C to 30°C) and pH (6.0 to 8.5) range
• Six-histidine sequence to facilitate its removal from the digested protein sample
• Greater than 85% single-band purity with no nonspecific protease contaminationApplications
Incubation with AcTEV™ Protease releases the protein of interest from the fusion tag. This is an effective way to remove solubility, secretion, detection, and purification tags from recombinant proteins.Enzyme specifications
Purified from E. coli expressing the AcTEV™ Protease gene.

Unit definition
One unit of AcTEV™ Protease cleaves 85% of a 3 µg control substrate in 1 hr at 30°C.

Unit reaction conditions
50 mM Tris-HCl (pH 8.0), 0.5 mM EDTA, 1 mM DTT, 3 µg control substrate, and 1 unit enzyme in 30 µL for 1 hr at 30°C. AcTEV™ Protease is functionally tested for the absence of any nonspecific protease activity.

WELQut Protease (5 U/µL) (Thermo Scientific™)

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.

Highlights

• Cleaves outside WELQ recognition sequence, without leaving additional amino acids bound to the target protein
• Highly specific to cognate recognition site, does not generate non-specific product bands, even after long incubation and using excess of protease
• Easy to remove from the reaction mixture using built-in His-tag
• Ideal for on-column proteolysis reactions

Applications

• Removal of N-terminal fusion tags from recombinant protein preparations.

Footnote

† 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.

SUMO Protease (Invitrogen™)

SUMO Protease, also known as Ulp, is a recombinant fragment of ULP1 (Ubl-specific protease 1) from Saccharomyces cerevisiae. It is highly specific for the SUMO protein fusion, recognizing the tertiary structure of SUMO rather than an amino acid sequence. The SUMO Protease features:

• Highly active cleavage
• No non-specific proteolysis (highly specific cleavage*)
• Activity from 2°C to 37°C
• A six-histidine sequence to facilitate its removal from the digested protein sample

Application: Removal of fusion tags from recombinant proteins.
Unit Definition: One unit of SUMO Protease is defined as the amount of enzyme needed to cleave 85% of 2µg of substrate protein at 30°C in one hour.
Quality Control: SUMO Protease has greater than 85% single-band purity with no non-specific protease contamination. It is functionally tested for the absence of any non-specific protease activity.
5 enzymes