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Protein splicing of a non‐canonical Clostridium thermocellum intein with N‐terminal Gln
Author(s) -
Powers Taryn L,
Drago Matthew J.,
Dorval Deirdre M.,
Connor Katherine R.,
Mills Kenneth V.
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a964-b
Subject(s) - intein , protein splicing , rna splicing , clostridium thermocellum , peptide bond , biochemistry , protein tag , chemistry , fusion protein , genetics , peptide , biology , gene , recombinant dna , enzyme , rna , cellulase
Protein splicing is a post‐translational modification by which an intervening polypeptide, called an intein, directs both its own excision from the flanking polypeptides, called exteins, and the ligation of the exteins by a peptide bond. The first step of the canonical mechanism of protein splicing is an amide to ester rearrangement of the peptide bond linking the N‐terminal extein and the first amino acid of the intein, which is a Ser or Cys. A small set of inteins have an N‐terminal Ala, and two such inteins have been shown to promote protein splicing, likely by bypassing the first step. We have examined the activity of an intein from Clostridium thermocellum that has an N‐terminal Gln. The intein was expressed as fusion protein between N‐ and C‐terminal affinity domains, and can facilitate protein splicing in vivo in E. coli . We have made site‐directed mutations at the N‐terminal position of the intein and at conserved, catalytic residues to examine the mechanism of splicing for this intein. This material is based upon work supported by the National Science Foundation (Grants DBI‐0320824 and MCB/CAREER‐0447647) and by the Research Corporation.