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Metabolic biotinylation of recombinant antibody by biotin ligase retained in the endoplasmic reticulum

Paper ID Volume ID Publish Year Pages File Format Full-Text
14149 1177 2007 9 PDF Available
Title
Metabolic biotinylation of recombinant antibody by biotin ligase retained in the endoplasmic reticulum
Abstract

Due to its strength and specificity, the interaction between avidin and biotin has been used in a variety of scientific and medical applications ranging from immunohistochemistry to drug targeting. The present study describes two methods for biotinylation of proteins secreted from eukaryotic cells using the Escherichia coli biotin protein ligase. In one system the biotin ligase was co-secreted from the cells along with substrate protein enabling extracellular biotinylation of the tagged protein. In the other system, biotin ligase was engineered to be retained in the endoplasmic reticulum (ER) and metabolically biotinylates the secretory protein as it passes through the ER. An engineered antibody fragment, a diabody with specificity for carcinoembryonic antigen (CEA) was fused to the biotin acceptor domain (123 amino acid) of Propionibacterium shermanii. Coexpression of the fusion protein with ER retained biotin ligase showed higher biotinylation efficiency than biotinylation by co-secreted ligase. Biotinylation of the anti-CEA diabody tagged with a short (15 amino acid, Biotin Avitag™) biotin acceptor peptide was also successful. Utilization of ER retained biotin ligase for biotinylation of protein is an attractive alternative for efficiently producing uniformly biotinylated recombinant proteins for a variety of avidin–biotin technologies.

Keywords
Biotinylation; Carcinoembryonic antigen; Engineered antibodies; Diabody; Biotin ligase; BirA
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Metabolic biotinylation of recombinant antibody by biotin ligase retained in the endoplasmic reticulum
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biomolecular Engineering - Volume 24, Issue 3, September 2007, Pages 283–291
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us