UGUS, a reporter for use with destabilizing N-termini |
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Authors: | D Zaccaria Roberta Greco H MacWilliams S Bozzaro A Ceccarelli |
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Affiliation: | Dipartimento di Scienze Cliniche e Biologiche, Università di Torino, Ospedale S.Luigi, 10043 Orbassano, Torino, Italy. |
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Abstract: | Due to constraints in vector construction, reporter polypeptides often carry N-terminal sequences of extraneous origin. Since protein half-life can be influenced by small determinants in the N-terminus, such foreign sequences can destabilize proteins and compromise results of reporter-based studies. We provide a real-life example of this problem (destabilizing sequences derived from a ribosomal protein) and show that it can be solved with the ubiquitin fusion technique, in which ubiquitin sequences are placed upstream of the reporter, in our case beta-glucuronidase. Post-translational processing by characterized pathways removes the ubiquitin together with destabilizing sequences, generating a stable reporter whose N-terminus is constant. |
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