Designed Protein Cages as Scaffolds for Building Multienzyme Materials

The functions of enzymes can be strongly affected by their higher-order spatial arrangements. In this study we combine multiple new technologiesdesigner protein cages and sortase-based enzymatic attachments between proteinsas a novel platform for organizing multiple enzymes (of one or more types)...

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Bibliographic Details
Published in:ACS synthetic biology 2020-02, Vol.9 (2), p.381-391
Main Authors: McConnell, Scott A, Cannon, Kevin A, Morgan, Christian, McAllister, Rachel, Amer, Brendan R, Clubb, Robert T, Yeates, Todd O
Format: Article
Language:English
Subjects:
Aminoacyltransferases - genetics
Aminoacyltransferases - metabolism
Bacterial Proteins - genetics
Bacterial Proteins - metabolism
Cellulase - genetics
Cellulase - metabolism
Cellulose - metabolism
Cysteine Endopeptidases - genetics
Cysteine Endopeptidases - metabolism
Nanocapsules - chemistry
Peptides - chemistry
Peptides - metabolism
Protein Engineering
Recombinant Proteins - biosynthesis
Recombinant Proteins - isolation & purification
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Summary:The functions of enzymes can be strongly affected by their higher-order spatial arrangements. In this study we combine multiple new technologiesdesigner protein cages and sortase-based enzymatic attachments between proteinsas a novel platform for organizing multiple enzymes (of one or more types) in specified configurations. As a scaffold we employ a previously characterized 24-subunit designed protein cage whose termini are outwardly exposed for attachment. As a first-use case, we test the attachment of two cellulase enzymes known to act synergistically in cellulose degradation. We show that, after endowing the termini of the cage subunits with a short “sort-tag” sequence (LPXTG) and the opposing termini of the cellulase enzymes with a short polyglycine sequence tag, addition of sortase covalently attaches the enzymes to the cage with good reactivity and high copy number. The doubly modified cages show enhanced activity in a cellulose degradation assay compared to enzymes in solution, and compared to a combination of singly modified cages. These new engineering strategies could be broadly useful in the development of enzymatic material and synthetic biology applications.
ISSN:2161-5063
2161-5063
DOI:10.1021/acssynbio.9b00407