With the rapid development of protein science, protein purification technology has an increasingly vital part in the field of biological research and biotechnology industry. As a core technique, it involves the integration of the coding information of the target protein into the host cells through genetic engineering methods, so that they can be induced to efficiently produce the required proteins. Subsequently, a series of sophisticated in vitro operation steps are carried out to achieve the high-purity extraction of proteins. Protein purification enables scientists to isolate single types of proteins, which is crucial for in-depth studies on the structure and function of proteins, development of new drugs, disease diagnosis and promotion of the application of biotechnology. This technology not only ensures the accuracy of scientific experiments but also continuously drives the development of life sciences.

Figure 1. Schematic diagram of affinity chromatography^[1]

In the field of protein engineering, fusion tags serve as a powerful tool, which can facilitate various experimental purposes by binding to the target proteins. The functions of common fusion tags are listed in the table below for reference. However, once these tags remain on the fusion proteins after fulfilling their initial auxiliary functions, they may have adverse effects on subsequent experiments, such as interfering with animal immunologic experiments or affecting the biological activity of proteins. Therefore, it is crucial to remove these unnecessary fusion tags to ensure the normal function of proteins and the accuracy of experiments.

Table 1. Introduction to the Functions of Common Fusion Tags and Enzyme Cleavage Methods

Fusion tag	Size (KDa)	Function	Common enzymatic cleavage methods
His	0.84	Beneficial for purification, able to purify soluble/inclusion body proteins.	TEV Protease
Flag	1.01	The purpose protein fused with Flag tag can be recognized by the antibody against Flag, so that the fusion protein containing Flag tag can be detected and identified by Western Blot, ELISA and other methods.	Enterokinase
GST	26	Enhance protein solubility, only able to purify soluble proteins and shield toxic proteins.	Thrombin
MBP	44.4	Enhance protein solubility and shield toxic proteins.	TEV Protease
NusA	55	Enhance protein solubility and shield toxic proteins.	Thrombin
SUMO	11.2	Enhance the solubility of proteins, promote protein from proteolytic hydrolysis, and improve the stability of proteins.	SUMO Protease

Today, we are delighted to introduce to you a cutting-edge product for the efficient and precise removal of fusion tags - UCF.ME^TM rTEV Protease. With its simple and easy-to-operate experimental procedure, this enzyme can conveniently cleave unnecessary tags from fusion proteins, thereby obtaining high-purity target proteins.

UCF.ME^TM rTEV Protease

TEV Protease is a protease widely used for the cleavage of recombinant protein fusion tags, known for its high site specificity. It strictly recognizes the seven-amino-acid sequence EXXYXQ↓(G/S) and performs precise cleavage between glutamine and glycine or serine. The most common seven-amino-acid sequence is Glu-Asn-Leu-Tyr-Phe-Gln↓-Gly. This specificity ensures the accuracy and efficiency in the protein processing procedure.

Figure 2. Schematic diagram of the action mechanism of TEV protease^[2]

UCF.ME^TM rTEV Protease is a recombinant protease that has been genetically engineered and finely purified. It not only retains the full functional activity of the natural TEV enzyme but also exhibits excellent stability and specificity over a wider temperature range. The host gDNA residue of this product is very low, ensuring higher cutting efficiency of protein fusion tags in practical applications and effectively reducing the risk of introducing exogenous substance residues. UCF.ME^TM rTEV Protease exhibits optimal activity under the conditions of pH 7.0 and 30°C. However, it maintains activity even under a wide range of conditions with pH 6.0-8.5 and temperature 4-30°C, to meet the different protein processing requirements. It is worth mentioning that UCF.ME^TM rTEV Protease has a 6×His tag at its N-terminus, which can be efficiently removed by Ni-NTA resin, thereby achieving precise purification of the target protein.

Performance display

1. High cleavage efficiency: The protein tag is cleaved more thoroughly

Using the fusion protein (3 μg) containing the UCF.ME^TM rTEV Protease cleavage site as the substrate, UCF.ME^TM rTEV Protease at 10, 5, and 3 U was added respectively, and the reaction was carried out at 30°C for 1 h. The cleavage effect was detected by agarose gel electrophoresis. The results showed that Yeasen's UCF.ME^TM rTEV Protease could cleave the substrate efficiently when the input amount was more than 3 U, with a cleavage efficiency of >90%.

Figure 3. Verification of the cleavage activity of UCF.ME^TM rTEV Protease

2. Low host gDNA residue: Effectively reduce the risk of introducing exogenous substance residues

By testing the host (E. coli) gDNA residues in different batches of UCF.ME^TM rTEV Protease, the results showed that the host gDNA residue of UCF.ME^TM rTEV Protease was far below <1 copy/U.

Figure 4. The results of host gDNA residue in UCF.ME^TM rTEV Protease

3. Wide application conditions: Able to meet different protein processing requirements.

By verifying the cleavage activity of UCF.ME^TM rTEV Protease under different conditions, the results showed that UCF.ME^TM rTEV Protease had strong activity under the conditions of 4-30°C, which could meet the different application requirements of customers.

Reaction time (h)	Cleavage activity under different temperatures (%)
Reaction time (h)	4℃	16℃	21℃	30℃
1	34	58	56	85
2	58	80	78	90
3	71	99	99	99
3.5	84	99	99	99

Yeasen's recommended fusion tag cutting protein products

Product Classification	Product Name	Product Number
TEV Protease	UCF.ME^TM rTEV Protease	20427ES
Enterokinase	Enterokinase, Recombinant	20395ES
SUMO Protease	SUMO Protease	20410ES
3C Protease	3C Protease	20409ES

References：

[1] Parikh I , Cuatrecasas P .Affinity Chromatography[J].Vox Sanguinis, 1972, 23(1-2):141-146.DOI:10.1159/000466530.

[2] Paththamperuma C, Page R C. Fluorescence dequenching assay for the activity of TEV protease[J]. Analytical biochemistry, 2022, 659: 114954.

UCF.METM rTEV Protease：An efficient solution for the precise and residue-free removal of protein tags