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Segmental isotopic labelling is a powerful method for the incorporation of stable isotopes into particular regions within proteins for NMR detection, thereby reducing the complexity of NMR spectra and offering the potential to perform sequential assignments. Here we have demonstrated segmental isotopic labelling of a domain in a multidomain protein both in(More)
BACKGROUND Protein trans-splicing by naturally occurring split DnaE inteins is used for protein ligation of foreign peptide fragments. In order to widen biotechnological applications of protein trans-splicing, it is highly desirable to have split inteins with shorter C-terminal fragments, which can be chemically synthesized. PRINCIPAL FINDINGS We report(More)
Segmental isotopic labeling is a powerful labeling technique for reducing nuclear magnetic resonance (NMR) signal overlap, which is associated with larger proteins by incorporating stable isotopes into only one region of a protein for NMR detections. Segmental isotopic labeling can not only reduce complexities of NMR spectra but also retain possibilities to(More)
Naturally split DnaE intein from Nostoc punctiforme (Npu) has robust protein trans-splicing activity and high tolerance of sequence variations at the splicing junctions. We determined the solution structure of a single chain variant of NpuDnaE intein by NMR spectroscopy. Based on the NMR structure and the backbone dynamics of the single chain NpuDnaE(More)
Alginate epimerases are large multidomain proteins capable of epimerising C5 on beta-D-mannuronic acid (M) turning it into alpha-L-guluronic acid (G) in a polymeric alginate. Azotobacter vinelandii secretes a family of seven epimerases, each of which is capable of producing alginates with characteristic G distribution patterns. All seven epimerases consist(More)
Protein splicing in trans by split inteins has increasingly become a powerful protein-engineering tool for protein ligation, both in vivo and in vitro. Over 100 naturally occurring and artificially engineered split inteins have been reported for protein ligation using protein trans-splicing. Here, we review the current status of the reported split inteins(More)
Protein splicing is an autocatalytic process involving self-excision of an internal protein domain, the intein, and concomitant ligation of the two flanking sequences, the exteins, with a peptide bond. Protein splicing can also take place in trans by naturally split inteins or artificially split inteins, ligating the exteins on two different polypeptide(More)
UNLABELLED Protein splicing in trans by split inteins has become a useful tool for protein engineering in vivo and in vitro. Inteins require Cys, Ser or Thr at the first residue of the C-terminal flanking sequence because a thiol or hydroxyl group in the side chains is a nucleophile indispensable for the trans-esterification step during protein splicing.(More)
Protein sequences are diversified on the DNA level by recombination and mutation and can be further increased on the RNA level by alternative RNA splicing, involving introns that have important roles in many biological processes. The protein version of introns (inteins), which catalyze protein splicing, were first reported in the 1990s. The biological roles(More)