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Proc Natl Acad Sci U S A. 2017 Aug 8;114(32):8538-8543. doi: 10.1073/pnas.1701083114. Epub 2017 Jul 24.

A promiscuous split intein with expanded protein engineering applications.

Author information

1
Department of Chemistry, Frick Laboratory, Princeton University, Princeton, NJ 08544.
2
Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461.
3
Department of Chemistry, Frick Laboratory, Princeton University, Princeton, NJ 08544; muir@princeton.edu.

Abstract

The protein trans-splicing (PTS) activity of naturally split inteins has found widespread use in chemical biology and biotechnology. However, currently used naturally split inteins suffer from an "extein dependence," whereby residues surrounding the splice junction strongly affect splicing efficiency, limiting the general applicability of many PTS-based methods. To address this, we describe a mechanism-guided protein engineering approach that imbues ultrafast DnaE split inteins with minimal extein dependence. The resulting "promiscuous" inteins are shown to be superior reagents for protein cyclization and protein semisynthesis, with the latter illustrated through the modification of native cellular chromatin. The promiscuous inteins reported here thus improve the applicability of existing PTS methods and should enable future efforts to engineer promiscuity into other naturally split inteins.

KEYWORDS:

chemical biology; intein splicing; protein engineering; protein semisynthesis

PMID:
28739907
PMCID:
PMC5559002
DOI:
10.1073/pnas.1701083114
[Indexed for MEDLINE]
Free PMC Article

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