期刊
ACS SYNTHETIC BIOLOGY
卷 9, 期 2, 页码 381-391出版社
AMER CHEMICAL SOC
DOI: 10.1021/acssynbio.9b00407
关键词
nanocages; sortase; cellulase; substrate channeling; bioconjugation
资金
- US Department of Energy Office of Science [DE-FC02-02ER63421]
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.
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