Amidohydrolases include glutamine PRPP amidotransferases (GATs), penicillin acylases/amidases (such as penicillin G acylases or PGAs and penicillin V acylases or PVAs), the 20S proteasome beta subunit (PRO), class II glutamine amidotransferases, glycosyl asparaginases (GAs), and also cephalosporin acylases, such as CAD, a class I cephalosporin acylase. The importance of Ntn hydrolases resides in the biosynthesis of beta-lactam antibiotics such as cephalosporin and penicillin. All enzymes in this superfamily share an unusual fold in which the nucleophile and other catalytic groups occupy equivalent sites. This fold provides both the capacity for nucleophilic attack and the possibility of autocatalytic processing. Posttranslational autoproteolysis is a mechanism used to activate many proteins via self-catalysed peptide bond rearrangements, which play an essential role in a wide variety of biological processes. They include activation cascades such as blood coagulation and fibrinolysis, cell death, embryonic development, protein targeting and degradation, viral protein processing, and zymogen activation.

A well-characterised enzyme containing this domain is CAD, a cephalosporin acylase from Pseudomonas diminuta. It consists of an alpha-subunit with 169 residues and a beta-subunit with 520 residues. The enzyme is described as a bowl-shaped molecule with two knobs attached to the bowl, facing each other. The A-knob contains mostly alpha-helices, whereas the B-knob contains mostly beta-sheets. The B-knob region is made up of seven beta-strands which form a distorted beta-barrel-like domain. This superfamily entry comprises structural domains similar to the beta-sheet B-knob region described in CAD.

Pfam clan [PfamClan:CL0052], Pfam family PFAM:PF01804, INTERPRO:IPR029055,PMID:7477383,PMID:11601852,PMID:11080627,PMID:16446446,PMID:23373797