The excision of an intervening protein sequence, an intein, from a precursor protein, with peptide bond formation between the flanking domains (the N-extein, originally attached to the N-terminal end of the intein, and the C-extein, originally attached to its C-terminal end) to produce the spliced protein. Intein sequences are homologous with homing endonucleases that transfer the intron gene to an intron-less allele, and some free intein proteins possess the endonuclease activity. The intein-C-extein interface is an asparagine-serine bond.

The splicing mechanism is autocatalytic, and is proposed to employ displacement of the N-extein from the intein by the β-hydroxy group of the serine. This produces a branched intermediate: to the C-extein serine residue are attached the intein in a peptide bond to the α-amino group and the N-extein in an ester bond to the β-hydroxy group. The asparagine residue cyclizes to free the intein, with a C-terminal aminosuccinimide residue formed from the asparagine; the N- and C-exteins remain joined by an ester bond. Displacement of the N-extein from the β-hydroxy group by serine's α-amino group produces the spliced protein. Some C-exteins have threonine or cysteine in place of the serine.