Engineering protein stability with an intra-molecular isopeptide bond

Show simple item record Young, Paul en Kwon, H en Baker, Edward en Squire, Christopher en
dc.coverage.spatial Canberra, Australia en 2016-08-09T22:55:13Z en 2016-04-04 en
dc.identifier.citation Synthetic Biology Cutting Edge Symposium, Canberra, Australia, 04 Apr 2016 - 05 Apr 2016. Synthetic Biology Cutting Edge Symposium. 04 Apr 2016 en
dc.identifier.uri en
dc.description.abstract Gram-positive bacterial pili and fimbriae are exposed to a hostile environment and must endure high tensile forces during the adhesion process. These ultra-stable proteins often contain internal crosslinks that impart crucial mechanical strength. Here, we report the first example of an isopeptide bond engineered de novo into an immunoglobulin-like protein domain. Through the site-directed mutagenesis of four residues we introduced isopeptide-forming residues into the hydrophobic core of FctB, an IgG-like minor pilin protein from Streptococcus pyogenes, to produce a covalent bond not present in the wild type protein. The isopeptide bond was engineered to covalently link the first and last β-strands, a position that is subjected to the highest mechanical stress. The presence of the engineered isopeptide-containing protein was confirmed by mass spectrometry and X-ray crystallography, and its thermal stability was measured as 81°C, an increase of 10°C over the wild type protein. This novel method for increasing the stability of IgG-like proteins has potential to be adopted by the field of antibody engineering, which share similar β-clasp Ig-type domains. en
dc.relation.ispartof Synthetic Biology Cutting Edge Symposium en
dc.rights Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher. en
dc.rights.uri en
dc.title Engineering protein stability with an intra-molecular isopeptide bond en
dc.type Conference Item en en
pubs.finish-date 2016-04-05 en
pubs.start-date 2016-04-04 en
dc.rights.accessrights en
pubs.subtype Abstract en
pubs.elements-id 529093 en Science en Biological Sciences en Science Research en Maurice Wilkins Centre (2010-2014) en
pubs.record-created-at-source-date 2016-05-31 en

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