Family N10

Family

Summary Holotypes Alignment Tree Genomes Structure Literature

Summary for family N10

Family type peptidaseN10.002 - intein-containing replicative DNA helicase precursor (Synechocystis sp. PCC 6803), MEROPS Accession MER0180503 (peptidase unit: 381-493, 644-810)
Content of familyFamily N10 includes self-cleaving proteins
History Identifier created: MEROPS 9.4 (31 January 2011)
An intein is a polypeptide insert into another protein (the extein) which is able to release itself from the host protein and splice the two portions of the extein together. Thus two proteins are generated from one: the extein and the intein. The intein is a multidomain protein, usually containing the domain responsible for cleavage and splicing divided into two portions, one at the N-terminus and one at the C-terminus, with an endonuclease domain known as a 'homing endonuclease' in the middle. The homing endonuclease recognizes a rare nucleotide sequence which it cleaves to leave a staggered break, usually of four bases. The staggered break allows the cleaved DNA to pair with the intein gene so that when the cell"s repair mechanism mend the break, the intein gene is used as a template with the effect that the intein gene is copied. The intein gene has replicated itself in a non-Mendelian manner, akin to that of some introns and transposons, and an intein gene can be considered an example of parasitic DNA. The burden on the host is light because the homing endonuclease recognition sequence is rare, and it is unusual for a genome to contain more than one intein, and because cleavage occurs within the recognition sequence, and once a gene has acquired an intein gene it cannot acquire another. Replication of the intein gene can occur during meiosis in eukaryote cells, and in prokaryote cells can occur by horizontal gene transfer (Raghavan & Minnick, 2009). The nature of the extein is irrelevent except that its gene must include the endonuclease recognition sequence. Although it is generally assumed that a protein can contain only one intein, there are several examples where two inteins can be identified in the protein sequence, and at least three proteins (from Methanocaldococcus and Thermococcus) that contain three potential inteins (and three potential homing endonucleases with the extein split into four fragments). There are also examples where the exteins and inteins are encoded by two open reading frames, where one gene encode the first part of the extein and part of the intein and the second encodes the remainder of the intein and the second part of the extein, an example being the DNA polymerase III subunit alpha from Synechocystis sp. PCC 6803 <%Martin et al., 2001[]%>.
Catalytic typeAsparagine
Active site residuesC381 N809 C,S,T810 
Active siteThree residues are involved in activity, the first (Cys, Ser or Thr) and last (Asn) residues of the intein, and the first residue of the second portion of the extein (Cys, Ser or Thr). Each of these acts as a nucleophile in each of the three reactions that occur to excise the intein and splice the remaining portions of the extein. The penultimate residue of the intein is often His, and this may be important for the cyclization of the Asn and thus cleavage of the bond between the Asn and the second portion of the extein. There are some inteins where the Asn is replaced by Gln. Gln cyclizes less readily than Asn, but a glutarimide intermediate can promote self-cleavage (Amitai et al., 2004). An example of an intein where the last residue is Gln is that from the DNA polymerase II large subunit of the archaean Haloarcula marismortui (N10.006).
Activities and specificitiesThe release of the intein from the extein is a self-cleaving event. Two peptide bond cleavages occur, at the residue immediately preceding the first residue of the intein and following the last residue. The first residue of the intein must by Cys, Ser or Thr and the last residue of the intein must be Asn or Gln. The first residue of the second portion of the extein must be Cys, Ser or Thr. The hydroxyl or thiol on the side chain of the first residue of the intein attacks the carbonyl carbon of the preceding amino acid to generate an ester or thiolester intermediate. Then transesterification occurs where the first residue of the second portion of the extein attacks the ester or thioester. This results in a branched intermediate where the first portion of the extein is transfered to the side chain of the first residue of the second portion and the peptide bond between the first portion of the extein and the first residue of the intein is broken. The branched intermediate thus has two N-termini. Then the last residue of the intein, which is usually Asn, cyclizes to form a succinimide ring, which results in the breakage of the peptide bond between the Asn and the first residue of the second portion of the extein. The intein is thus released. The ester or thioester bond in the extein then rapidly undergoes an acyl rearrangement to form a normal peptide bond, which is thermodynamically more stable (Raghavan & Minnick, 2009). Neither peptide bond cleavage involves hydrolysis, and hydrolysis, if it occurs, converts the succinimide ring back to Asn.
Molecular structureSeveral crystal structures have been solved, including the intein-containing RecA protein from Mycobacterium tuberculosis (N10.002; Datta et al., 2003) and the inteins from the DNA polymerase III subunit alpha precursor from Synechocystis sp. PCC 6803 (N10.002; Sun et al., 2005), DNA-dependent DNA polymerase family B protein from Thermococcus kodakarensis (N10.007; Matsumara et al., 2006)and DNA gyrase subunit A from Mycobacterium xenopi (Klabunde et al., 1998). The intein structures are disc-like with the active site residues in the centre. The fold is similar to that of the self-cleaving hedgehog proteins (family C46) hence both families are included in the same clan.
ClanPD
SubclanPD(N)
Distribution of family Bacteria details  
Archaea details  
Protozoa -  
Fungi -  
Plants -  
Animals -  
Viruses details  
Biological functions
Statistics for family N10Sequences:458
Identifiers:9
Identifiers with PDB entries:7
Downloadable files Sequence library (FastA format)
Sequence alignment (FastA format)
Phylogenetic tree (Newick format)
Peptidases and Homologues MEROPS ID Structure
intein-containing DNA gyrase subunit A precursorN10.001-
intein-containing replicative DNA helicase precursorN10.002Yes
intein-containing DNA polymerase III subunit alpha precursorN10.003Yes
intein-containing translation initiation factor IF-2 precursorN10.004-
intein-containing DNA polymerase II large subunit DP2 precursor Mername-AA281N10.005Yes
intein-containing DNA polymerase II large subunit DP2 precursor Mername-AA282N10.006-
intein-containing DNA-dependent DNA polymerase precursorN10.007Yes
intein-containing DNA gyrase subunit A (Mycobacterium xenopi)N10.008Yes
Mtu recA intein (Mycobacterium sp.)N10.009Yes
family N10 non-lyase homologuenon-peptidase homologue-
family N10 unassigned peptide lyaseunassignedYes