Conserved sequence features of inteins (protein introns) and their use in identifying new inteins and related proteins

Abstract

Inteins (protein introns) are internal portions of protein sequences that are posttranslationally excised while the flanking regions are spliced together, making an additional protein product. Inteins have been found in a number of homologous genes in yeast, mycobacteria, and extreme thermophile archaebacteria. The inteins are probably multifunctional, autocatalyzing their own splicing, and some were also shown to be DNA endonucleases. The splice junction regions and two regions similar to homing endonucleases were thought to be the only common sequence features of inteins.This work analyzed all published intein sequences with recently developed methods for detecting weak, conserved sequence features. The methods complemented each other in the identification and assessment of several patterns characterizing the intein sequences. New intein conserved features are discovered and the known ones are quantitatively described and localized. The general sequence description of all the known inteins is derived from the motifs and their relative positions. The intein sequence description is used to search the sequence databases for intein-like proteins. A sequence region in a mycobacterial open reading frame possessing all of the intein motifs and absent from sequences homologous to both of its flanking sequences is identified as an intein. A newly discovered putative intein in red algae chloroplasts is found not to contain the endonuclease motifs present in all other inteins. The yeast HO endonuclease is found to have an overall intein-like structure and a few viral polyprotein cleavage sites are found to be significantly similar to the inteins amino-end splice junction motif. The intein features described may serve for detection of intein sequences.

The Deinococcus radiodurans Snf2 intein caught in the act: Detection of the Class 3 intein signature Block F branched intermediate

“…5 Asp is not commonly found at B:7 although it is present in a few other inteins. [1][2][3][4][5] Mutation of Asp61 to Ala inhibited reactions at the N-terminal splice junction reducing the amount of spliced products and increasing the amount of C-terminal cleavage products [ Fig. 4(B) and Table II].…”

“…1). [2][3][4][5] Intein residues are referred to using the Block letter and the Block position separated by a colon.…”

“…1). [1][2][3][4][5] Two variations in the intein-mediated protein splicing mechanism were previously described, which involve direct attack on an amide bond at the N-terminal splice junction to generate a BI. [5][6][7]12,13 In Class 2 inteins, exemplified by the Methanococcus jannaschii (Mja) KlbA intein, 12,13 the Block G BI (III) is formed by nucleophilic attack of Cysþ1 on the peptide bond at the N-terminal splice junction (Fig.…”

“…1) and the mechanistic roles that they play. [1][2][3][4][5][6][7] Putative Class 3 inteins can be identified by class-specific sequence features: (a) the absence of a Cys, Ser, or Thr N-terminal nucleophile at A:1, (b) a noncontiguous WCT triplet at B:12, F:4, and G:5, and (c) Thrþ1 or Serþ1 as the first C-extein residue at G:8 instead of the more common Cysþ1 (Fig. 1).…”

“…Conserved positions are populated by groups of residues that have similar properties except for the Block B His (B:10), which is almost invariant. [1][2][3][4][5] An exemplary sequence is shown for each intein class. The most common residue found at selected positions is underlined and the common variations for these positions are shown.…”