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Controlling Macromolecular Topology with Genetically Encoded SpyTag-SpyCatcher Chemistry

Zhang, Wen-Bin and Sun, Fei and Tirrell, David A. and Arnold, Frances H. (2013) Controlling Macromolecular Topology with Genetically Encoded SpyTag-SpyCatcher Chemistry. Journal of the American Chemical Society, 135 (37). pp. 13988-13997. ISSN 0002-7863. doi:10.1021/ja4076452.

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Control of molecular topology constitutes a fundamental challenge in macromolecular chemistry. Here we describe the synthesis and characterization of artificial elastin-like proteins (ELPs) with unconventional nonlinear topologies including circular, tadpole, star, and H-shaped proteins using genetically encoded SpyTag–SpyCatcher chemistry. SpyTag is a short polypeptide that binds its protein partner SpyCatcher and forms isopeptide bonds under physiological conditions. Sequences encoding SpyTag and SpyCatcher can be strategically placed into ELP genes to direct post-translational topological modification in situ. Placement of SpyTag at the N-terminus and SpyCatcher at the C-terminus directs formation of circular ELPs. Induction of expression at 16 °C with 10 μM IPTG yields 80% monomeric cyclic protein. When SpyTag is placed in the middle of the chain, it exhibits an even stronger tendency toward cyclization, yielding up to 94% monomeric tadpole proteins. Telechelic ELPs containing either SpyTag or SpyCatcher can be expressed, purified, and then coupled spontaneously upon mixing in vitro. Block proteins, 3-arm or 4-arm star proteins, and H-shaped proteins have been prepared, with the folded CnaB2 domain that results from the SpyTag–SpyCatcher reaction as the molecular core or branch junction. The modular character of the SpyTag–SpyCatcher strategy should make it useful for preparing nonlinear macromolecules of diverse sequence and structure.

Item Type:Article
Related URLs:
URLURL TypeDescription DOIArticle
Tirrell, David A.0000-0003-3175-4596
Arnold, Frances H.0000-0002-4027-364X
Additional Information:© 2013 American Chemical Society. Received: July 24, 2013; Published: August 21, 2013. This research was funded by National Science Foundation (DMR 1206121) and by the Gordon and Betty Moore Foundation through Grant GBMF2809 to the Caltech Programmable Molecular Technology Initiative.
Funding AgencyGrant Number
Gordon and Betty Moore FoundationGBMF2809
Issue or Number:37
Record Number:CaltechAUTHORS:20140221-084933853
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Official Citation:Controlling Macromolecular Topology with Genetically Encoded SpyTag–SpyCatcher Chemistry Wen-Bin Zhang, Fei Sun, David A. Tirrell, and Frances H. Arnold Journal of the American Chemical Society 2013 135 (37), 13988-13997
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:43920
Deposited By: Tony Diaz
Deposited On:13 Mar 2014 22:59
Last Modified:10 Nov 2021 16:45

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