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ARS Home » Pacific West Area » Albany, California » Western Regional Research Center » Healthy Processed Foods Research » Research » Publications at this Location » Publication #333803
Research Project: Defining, Measuring, and Mitigating Attributes that Adversely Impact the Quality and Marketability of Foods

Location: Healthy Processed Foods Research

Title: A mutant sumo facilitates quick plasmid construction for expressing proteins with native N-termini after fusion tag removal

Author
item Zhang, Yuzhu
item FAN, YUTING - Jiangnan University

Submitted to: Journal of Molecular Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/30/2017
Publication Date: 3/27/2017
Citation: Zhang, Y., Fan, Y. 2017. A mutant sumo facilitates quick plasmid construction for expressing proteins with native N-termini after fusion tag removal. Journal of Molecular Biology. 59(4-5):157-167. https://doi.org/10.1007/s12033-017-9998-6.
DOI: https://doi.org/10.1007/s12033-017-9998-6

Interpretive Summary: IgE antibodies are produced to both linear and conformational epitopes. For understanding the allergenicity of food allergens, characterizing both linear and conformational epitopes is required. For such studies, a relatively large amount of highly purified recombinant food allergens are often require. Sumo was successfully used as a peptidase recognition site to produce a recombinant food allergen with a native N-terminus. The cloning vector generated in this study can be used for quick generation of plasmid for producing food recombinant food allergens for studying the importance of conformational IgE epitopes.

Technical Abstract: Sumo is one of the fusion tags commonly used to enhance the solubility and yield of recombinant proteins. One advantage of using sumo is that the removal of the sumo tag is highly specific because its recognition by the ULP sumo protease is determined by its structural characteristics, instead of the sequence of a short peptide. Recently, we have shown that sumo can also be used as protease recognition site to facilitate the removal of other fusion tags, such as MBP, when sumo itself is not good enough to enhance the solubility of the target protein. Using sumo as a recognition site is highly desirable when the target protein needs to have its native N-terminus. However, constructing such a plasmid involved more than common cloning steps due the lack of multiple cloning sites within the available vector. Here, we report the construction of a sumo mutant which did not change its recognition and cleavage by ULP but enabled the inclusion of Pvu II site near the 3’ end of the sumo coding sequence. Because many food allergens derived from plant sources have non-met N-termini due to the removal of signal peptides, the new construct is especially useful in the expression of recombinant food allergens for studying the importance of conformational IgE epitopes.