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Tagging and Fusion Proteins

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Proteins are involved in several cellular processes and biochemical reactions. Analyzing a specific protein of interest requires it to be isolated from the other proteins in the cell. This is achieved by overexpressing the specific gene in a suitable host to produce large quantities of the target protein. A tag or label is recombined with the gene to produce a fusion protein containing the target protein and the tag. The tags on these fusion proteins can then be used for easy detection and...

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Related Experiment Video

Updated: Apr 9, 2026

Protease- and Acid-catalyzed Labeling Workflows Employing 18O-enriched Water

Protease- and Acid-catalyzed Labeling Workflows Employing 18O-enriched Water

Published on: February 20, 2013

Proteolytic Affinity Tag Cleavage.

Helena Block¹, Barbara Maertens¹, Anne Spriestersbach¹

¹QIAGEN GmbH, Research and Development, Qiagenstrasse 1, 40724 Hilden, Germany.

Methods in Enzymology

|June 23, 2015

Summary

This summary is machine-generated.

This study introduces two protease systems, TAGZyme and Factor Xa, for efficient protein purification. These methods ensure target proteins are free from vector-encoded amino acids and purification tags, simplifying downstream applications.

Keywords:

DAPase enzyme Exoproteolytic cleavage GST fusion protein Glutathione S-Transferase tag (GST tag)Immobilized metal ion affinity chromatography (IMAC)Qcyclase enzymes TAGZyme Xa removal resin

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Area of Science:

Biochemistry
Molecular Biology
Protein Chemistry

Background:

Protein purification often results in residual amino acids or tags, complicating subsequent experiments.
Efficient methods are needed to remove protease recognition sites and purification tags from recombinant proteins.

Purpose of the Study:

To present protocols for using the TAGZyme (dipeptidyl-aminopeptidase-1, DPP1) and Factor Xa protease systems.
To demonstrate the recovery of target proteins free of vector-encoded amino acids and protease recognition sites.

Main Methods:

Utilized the dipeptidyl-aminopeptidase-1 (DPP1, DAPase) exoprotease-based TAGZyme system.
Employed the endoprotease, Factor Xa.
Applied subtractive chromatography for protease removal.
Used Immobilized Metal Ion Affinity Chromatography (IMAC) for TAGZyme enzyme removal.
Employed Xa Removal Resin for Factor Xa removal.

Main Results:

Achieved recovery of target proteins entirely free of vector-encoded amino acids.
Successfully removed protease recognition sites from the target protein preparations.
Demonstrated efficient removal of both TAGZyme and Factor Xa proteases via subtractive chromatography.

Conclusions:

The TAGZyme and Factor Xa systems provide effective strategies for producing highly pure recombinant proteins.
These methods simplify protein purification by enabling the removal of unwanted amino acids and purification tags.
The described protocols offer robust solutions for researchers requiring tag-free proteins.