Traceless protein semi-synthesis in cells using the promiscuous ultra-fast split intein NrdJ-1

Area of Science:

• Biochemistry
• Molecular Biology
• Chemical Biology

Background:

• Chemistry-driven protein manipulation offers advantages beyond standard genetics for biomedical research.
• Intein-mediated protein splicing is a powerful platform for both in vitro and in vivo applications.
• Split inteins enable protein ligation and modification by reconstituting functional inteins from separate fragments.

Purpose of the Study:

• To provide a detailed biochemical characterization of the NrdJ-1 split intein.
• To assess the efficiency and kinetics of NrdJ-1 mediated protein splicing.
• To demonstrate the utility of NrdJ-1 for generating chemically modified proteins and traceless protein editing.

Main Methods:

• Biochemical assays to characterize NrdJ-1 split intein activity.
• Analysis of splicing kinetics and dependence on sequence context.
• Application of NrdJ-1 for in vitro and in vivo protein modification, including generation of semisynthetic chromatin.

Main Results:

• NrdJ-1 exhibits rapid splicing kinetics largely independent of the splice junction sequence.
• The split intein demonstrates high efficiency and promiscuity in protein splicing.
• Successful traceless generation of semisynthetic chromatin with post-translational modifications was achieved using NrdJ-1.

Conclusions:

• NrdJ-1 is a highly efficient and versatile split intein for protein editing.
• Its rapid and context-independent splicing makes it suitable for diverse in vitro and in vivo applications.
• NrdJ-1 represents a superior choice for intracellular biochemical applications requiring traceless protein modification.