EC 3. Hydrolases
These enzymes catalyse the hydrolysis of various bonds. Some of these enzymes pose problems because they have a very wide specificity, and it is not easy to decide if two preparations described by different authors are the same, or if they should be listed under different entries. While the systematic name always includes 'hydrolase', the Accepted name is, in most cases, formed by the name of the substrate with the suffix -ase. It is understood that the name of the substrate with this suffix, and no other indicator, means a hydrolytic enzyme.
EC 3.1 Acting on ester bonds
The esterases are subdivided into those acting on carboxylic esters (EC 3.1.1), thioester hydrolases (EC 3.1.2), phosphoric monoester hydrolases, the phosphatases (EC 3.1.3), phosphodiester hydrolases (EC 3.1.4), triphosphoric monoester hydrolases (EC 3.1.5), sulfatases (EC 3.1.6), diphosphoric monoesterases (EC 3.1.7) and phosphoric triester hydrolases (EC 3.1.8). The nucleases, previously included under EC 3.1.4, are now placed in a number of new sub-subclasses: the exonucleases (EC 3.1.11-16), and the endonucleases (EC 3.1.21-31).
EC 3.1.23 and EC 3.1.24EC 3.2 Glycosylases
In a previous edition, site specific endodeoxyribonucleases were set out individually here as subclasses EC 3.1.23 and EC 3.1.24, with 113 separate entries. These are now included in three entries EC 220.127.116.11, EC 18.104.22.168 and EC 22.214.171.124. A complete listing of all of these enzymes has been produced by R.J. Roberts and is available at http://rebase.neb.com/rebase/rebase.html.
Glycosylases are classified under hydrolases, although some of them can also transfer glycosyl residues to oligosaccharides, polysaccharides and other alcoholic acceptors. The glycosylases are subdivided into those hydrolysing O- or S-glycosyl compounds (EC 3.2.1 and EC 3.2.3), i.e. glycosides, and those hydrolysing N-glycosyl compounds (EC 3.2.2). Accepted names for enzymes acting on D-sugars or their derivatives do not contain 'D', unless ambiguity would result from the common existance of the corresponding L-sugar.
EC 3.3 Acting on ether bonds
To this small subclass belong enzymes acting on ether bonds. It is subdivided into those hydrolysing on thioether and trialkylsulfonium compounds (EC 3.3.1) and those acting on ethers (EC 3.3.2).
EC 3.4 Acting on Peptide Bonds (Peptidases)
It is recommended that the term "peptidase" be used as synonymous with "peptide hydrolase" for any enzyme that hydrolyses peptide bonds. Peptidases are recommended to be further divided into "exopeptidases" that act only near a terminus of a polypeptide chain and "endopeptidases" that act internally in polypeptide chains. The types of exopeptidases and endopeptidases are described more fully below. The usage of "peptidase" now recommended is synonymous with "protease" as it was originally used  as a general term for both exopeptidases and endopeptidases, but it should be noted that previously, in Enzyme Nomenclature (1984), "peptidase" was restricted to the enzymes included in sub-subclasses EC 3.4.11-19, the exopeptidases. Also, the term "proteinase" used previously for the enzymes included in sub-subclasses EC 3.4.21-99 carried the same meaning as "endopeptidase", and has been replaced by "endopeptidase" for consistency.
The nomenclature of the peptidases is troublesome. Their specificity is commonly difficult to define, depending upon the nature of several amino acid residues around the peptide bond to be hydrolysed and also on the conformation of the substrate polypeptide chain. A classification involving the additional criterion of catalytic mechanism is therefore used.
Two sets of sub-subclasses of peptidases are recognised, those of the exopeptidases (EC 3.4.11-19) and those of the endopeptidases (EC 3.4.21-24 and EC 3.4.99). The exopeptidases act only near the ends of polypeptide chains, and those acting at a free N-terminus liberate a single amino-acid residue (aminopeptidases, EC 3.4.11), or a dipeptide or a tripeptide (dipeptidyl-peptidases and tripeptidyl-peptidases, EC 3.4.14). The exopeptidases acting at a free C-terminus liberate a single residue (carboxypeptidases, EC 3.4.16-18) or a dipeptide (peptidyl-dipeptidases, EC 3.4.15). The carboxypeptidases are allocated to four groups on the basis of catalytic mechanism: the serine-type carboxypeptidases (EC 3.4.16), the metallocarboxypeptidases (EC 3.4.17) and the cysteine-type carboxypeptidases (EC 3.4.18). Other exopeptidases are specific for dipeptides (dipeptidases, EC 3.4.13), or remove terminal residues that are substituted, cyclized or linked by isopeptide bonds (peptide linkages other than those of α-carboxyl to α-amino groups) (omega peptidases, EC 3.4.19).
The endopeptidases are divided into sub-subclasses on the basis of catalytic mechanism, and specificity is used only to identify individual enzymes within the groups. These are the sub-subclasses of serine endopeptidases (EC 3.4.21), cysteine endopeptidases (EC 3.4.22), aspartic endopeptidases (EC 3.4.23), metalloendopeptidases (EC 3.4.24) and threonine endopeptidases (EC 3.4.25). Endopeptidases that could not be assigned to any of the sub-subclasses EC 3.4.21-25 were listed in sub-subclass EC 3.4.99.
There are characteristic inhibitors of the members of each catalytic type of endopeptidase; to save space these have not been listed separately for each individual enzyme but are reviewed in  and . A general source of information on peptidases that similarly has not been cited for each individual enzyme is reference .
In describing the specificity of peptidases, use is made of a model in which the catalytic site is considered to be flanked on one or both sides by specificity subsites, each able to accommodate the sidechain of a single amino acid residue (based on ). These sites are numbered from the catalytic site, S1...Sn towards the N-terminus of the substrate, and S1'...Sn' towards the C-terminus. The residues they accommodate are numbered P1...Pn, and P1'...Pn', respectively, as follows:
Substrate: - P3 - P2 - P1 P1'- P2'- P3'-
Enzyme: - S3 - S2 - S1 * S1'- S2'- S3'-
In this representation, the catalytic site of the enzyme is marked *. The peptide bond cleaved (the scissile bond) is indicated by the symbol '' or a hyphen in the structural formula of the substrate, or a hyphen in the name of the enzyme. In describing the specificity of endopeptidases, the term "oligopeptidase" is used to refer to those that act only on substrates smaller than proteins.
Finally, in describing the specificity of endopeptidases, the term oligopeptidase' is used to refer to those that act optimally on substrates smaller than proteins.
Families of peptidases are referred to by use of the numbering system of Rawlings & Barrett [6, 7].
1. Grassmann, W. & Dyckerhoff, H. Über die Proteinase und die Polypeptidase der Hefe. 13. Abhandlung über Pflanzenproteasen in der von R. Willstätter und Mitarbeitern begonnenen Untersuchungsreihe. Hoppe-Seyler's Z. Physiol. Chem. 179 (1928) 41-78.
2. Barrett, A. J. & Salvesen, G. S. (eds) Proteinase Inhibitors, Elsevier Science Publishers, Amsterdam (1986).
3. Beynon, R. J. & Bond, J. S. Proteolytic Enzymes. A Practical Approach. IRL Press, Oxford (1989).
4. Barrett, A. J., Rawlings, N. D. & Woessner, J. F. (eds) Handbook of Proteolytic Enzymes, Academic Press, London (1998).
5. Berger, A. and Schechter, I. Mapping the active site of papain with the aid of peptide substrates and inhibitors. Philos. Trans. R. Soc. London, Ser. B:Biol. Sci. 257 (1970) 249-264.
6. Rawlings, N.D. and Barrett, A.J. In: Methods Enzymol. 244 (1994) 19-61 and 461-486; Methods Enzymol. 248 (1995) 105-120 and 183-228.
7. Rawlings, N. D. and Barrett, A. J. MEROPS: the peptidase database. Nucleic Acids Research 27 (1999) 325-331.
EC 3.5 Acting on Carbon-Nitrogen Bonds, other than Peptide Bonds
To this subclass belong those enzymes hydrolysing amides, amidines and other C-N bonds. The sub-subclasses are separated on the basis of the substrate: linear amides (EC 3.5.1), cyclic amides (EC 3.5.2), linear amidines (EC 3.5.3), cyclic amidines (EC 3.5.4), nitriles (EC 3.5.5) and other compounds (EC 3.5.99).
EC 3.6 Acting on Acid Anhydrides
To this subclass belong mainly the enzymes acting on diphosphate bonds in compounds such as nucleoside di- and tri-phosphates (EC 3.6.1), on sulfonyl-containing anhydrides such as adenylylsulfate (EC 3.6.2) and on acid anhydrides; catalysing transmembrane movement of substances (EC 3.6.3).
EC 3.6.3 Acting on acid anhydrides; catalysing transmembrane movement of substances.EC 3.7 Acting on Carbon-Carbon Bonds
Several types of ATP phosphohydrolase are listed here. Entries EC 126.96.36.199 to EC 188.8.131.52 and EC 184.108.40.206 are enzymes undergoing covalent phosphorylation of an aspartate residue during the transport cycle; entries EC 220.127.116.11 and EC 18.104.22.168 refer to enzymes of complicated membrane and non-membrane location that can also serve in ATP synthesis; entry EC 22.214.171.124 is a multisubunit enzyme that is involved in arsenite transport only; entries EC 126.96.36.199 to EC 188.8.131.52 are two-domain enzymes of the ABC family; entries EC 184.108.40.206 and EC 220.127.116.11 are parts of a complex protein-transporting machinery in mitochondria and chloroplasts.
There are relatively few carbon-carbon hydrolases; they mostly catalyse the hydrolysis of 3-oxo-carboxylic acids.
EC 3.8 Acting on Halide Bonds
These enzymes hydrolyse organic halides.
EC 3.9 Acting on Phosphorus-Nitrogen Bonds
This is the phosphoamidase group.
EC 3.10 Acting on Sulfur-Nitrogen Bonds
EC 3.11 Acting on Carbon-Phosphorus Bonds
The enzymes in this subclass hydrolyse C-phosphono-groups
EC 3.12 Acting on Sulfur-Sulfur Bonds
EC 3.13 Acting on Carbon-Sulfur Bonds
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