Serine peptidase

Du/tripeptidyl-peptidases Peptidyl-dipeptidases Serine carboxypeptidases MetaUocarboxypeptidases Cysteine carboxypeptidases Omega peptidases Serine endopeptidases Cysteine endopeptidases Aspartic endopeptidases MetaUoendopeptidases Threonine endopeptidases Other endopeptidases... 

Serine peptidases, serine proteases the most studied class of peptidases. They have a reactive serine residue, for example, the hydrolysis of a peptide substrate involves an acyl enzyme intermediate in which the hydroxyl group of Ser (chymotrypsin numbering system) is acylated by the acyl moiety of the substrate, thus releasing the amine fragment of the substrate as the first product. The formation of the acyl enzyme is the slow step in peptide bond hydrolysis, but acylenzyme often accumulates in the hydrolysis of ester substrates. The acyl enzyme thus formed will be the same for a series of substrates which differ in their leaving group. The active site of serine peptidases is complementary in stmc-ture to the transition state of the reaction, a structure which is very close to the tetrahedral adduct of Ser and the carbonyl... 

Peptidase Serine RONOZYME ProAct (Novozyme) Genetically modified strain of B. licheniformis Animal feed Caloni (2009)... 

Peptidase Serine (subtiiisins) Alcalase (Novozyme) B. licheniformis Detergent Maurer (2004), Saeki et al. (2007)... 

Several drugs in current medical use are mechanism-based enzyme inactivators. Eor example, the antibiotic penicillin exerts its effects by covalently reacting with an essential serine residue in the active site of glycoprotein peptidase, an enzyme that acts to cross-link the peptidoglycan chains during synthesis of bacterial cell walls (Eigure 14.17). Once cell wall synthesis is blocked, the bacterial cells are very susceptible to rupture by osmotic lysis, and bacterial growth is halted. 

For many serine and cysteine peptidases catalysis first involves formation of a complex known as an acyl intermediate. An essential residue is required to stabilize this intermediate by helping to form the oxyanion hole. In cathepsin B a glutamine performs this role and sometimes a catalytic tetrad (Gin, Cys, His, Asn) is referred too. In chymotrypsin, a glycine is essential for stabilizing the oxyanion hole. 

Threonine peptidases (and some cysteine and serine peptidases) have only one active site residue, which is the N-terminus of the mature protein. Such a peptidase is known as an N-terminal nucleophile hydrolase or Ntn-hydrolase. The amino group of the N-terminal residue performs the role of the general base. The catalytic subunits of the proteasome are examples of Ntn-hydrolases. 

Peptidases have been classified by the MEROPS system since 1993 [2], which has been available viatheMEROPS database since 1996 [3]. The classification is based on sequence and structural similarities. Because peptidases are often multidomain proteins, only the domain directly involved in catalysis, and which beais the active site residues, is used in comparisons. This domain is known as the peptidase unit. Peptidases with statistically significant peptidase unit sequence similarities are included in the same family. To date 186 families of peptidase have been detected. Examples from 86 of these families are known in humans. A family is named from a letter representing the catalytic type ( A for aspartic, G for glutamic, M for metallo, C for cysteine, S for serine and T for threonine) plus a number. Examples of family names are shown in Table 1. There are 53 families of metallopeptidases (24 in human), 14 of aspartic peptidases (three of which are found in human), 62 of cysteine peptidases (19 in human), 42 of serine peptidases (17 in human), four of threonine peptidases (three in human), one of ghitamicpeptidases and nine families for which the catalytic type is unknown (one in human). It should be noted that within a family not all of the members will be peptidases. Usually non-peptidase homologues are a minority and can be easily detected because not all of the active site residues are conserved. 

All peptidases within a family will have a similar tertiary structure, and it is not uncommon for peptidases in one family to have a similar structure to peptidases in another family, even though there is no significant sequence similarity. Families of peptidases with similar structures and the same order of active site residues are included in the same clan. A clan name consists of two letters, the first representing the catalytic type as before, but with the extra letter P , and the second assigned sequentially. Unlike families, a clan may contain peptidases of more than one catalytic type. So far this has only been seen for peptidases with protein nucleophiles, and these clans are named with an initial P . Only three such clans are known. Clan PA includes peptidases with a chymotrypsin-like fold, which besides serine peptidases such as chymotrypsin... 

It is recommended that a well-characterized peptidase should have a trivial name. Although not rigidly adhered to, there is a different suffix for each catalytic type, metallopeptidases names end with lysin , aspartic peptidases with pepsin , cysteine pqrtidase with ain and serine peptidases with in . 

The action of a peptidase can be neutralized by an inhibitor. Some inhibitors are very broad in their action and are capable of inhibiting many different peptidases, including peptidases of different catalytic types. Some inhibitors are assumed to be specific for a particular catalytic type, but can inhibit peptidases of different types. Leupeptin, for example, is widely used as an inhibitor of serine peptidases from family SI, but it is also known to inhibit cysteine peptidases from family Cl. Cysteine pqrtidase inhibitors such as iodoacetic acid interact with the thiol of the catalytic cysteine. However, this reduction can occur on any thiol group and can affect other, predominantly intracellular, peptidases with a thiol dependency. One example is thimet oligopepti-dase. Metal chelators such as EDTA can inhibit meta-llopeptidases, but can also affect peptidases that have a requirement for metal ions that is indq>endent of their catalytic activity, such as the calcium-dependent cysteine endopqrtidase calpain 1. 

Inhibitors which interact only with peptidases of one catalytic type include pepstatin (aspartic peptidases) E64 (cysteine peptidases from clan CA) diisopropyl fluorophosphates (DFP) and phenylmethane sulfonyl-fluoride (PMSF) (serine peptidases). Bestatin is a useful inhibitor of aminopeptidases. 

The antibiotic activity of certain (3-lactams depends largely on their interaction with two different groups of bacterial enzymes. (3-Lactams, like the penicillins and cephalosporins, inhibit the DD-peptidases/transpeptidases that are responsible for the final step of bacterial cell wall biosynthesis.63 Unfortunately, they are themselves destroyed by the [3-lactamases,64 which thereby provide much of the resistance to these antibiotics. Class A, C, and D [3-lactamases and DD-peptidases all have a conserved serine residue in the active site whose hydroxyl group is the primary nucleophile that attacks the substrate carbonyl. Catalysis in both cases involves a double-displacement reaction with the transient formation of an acyl-enzyme intermediate. The major distinction between [3-lactamases and their evolutionary parents the DD-peptidase residues is the lifetime of the acyl-enzyme it is short in (3-lactamases and long in the DD-peptidases.65-67... 

The functionalized phenaceturates 16 (Fig. 11.10) are substrates of class A and C [3-lactamases, especially the class C enzymes, as observed with the parent unfunctionalized phenaceturates 15. They are also modest inhibitors of these enzymes and the serine DD-peptidase of Streptomyces R61. The inhibition of class C [3-lactamases is turnover dependent, as expected for a mechanism-based inhibitor. Inhibition is not very dependent on the nature of the leaving group, suggesting that the QM is generated in solution after the product phenol has been released from the active site. It therefore... 

Rawlings, N. D. Barrett, A. J. Introduction serine peptidases and their clans. In Handbook of Proteolytic Enzymes, 2nd edition Barrett, A. J. Rawlings, N. D. Woessner, J. F., Eds. Elsevier Academic Press Amsterdam, Boston, Heidelberg, London, New York, Oxford, Paris, San Diego, San Francisco, Singapore, Sydney, Tokyo 2004, pp 1417-1439. 

Volume 244. Proteolytic Enzymes Serine and Cysteine Peptidases Edited by Alan J. Barrett... 

This is a 29-kDa protein that has NH 2-terminal sequence homology with elastase and cathepsin G. However, it contains glycine and not serine at the predicted catalytic site, and so lacks protease and peptidase activity. Purified azurocidin kills a range of organisms (e.g. E. coli, S.faecalis, and C. albicans) in vitro. It functions optimally at pH 5.5 and in conditions of low ionic strength. 

Its length can vary from 30 to more than 100 residues. It is rich in serine and threonine but deficient in acidic residues. It is not clear whether it has the three-domain structure like the signal peptide (von Heijne et al., 1989). Recently, a chloroplast-processing enzyme was identified as the general stromal processing peptidase (Richter and Lamppa, 1998). 

The peptidases were separated into catalytic types according to the chemical nature of the group responsible for catalysis. The major catalytic types are, thus, Serine (and the related Threonine), Cysteine, Aspartic, Metallo, and As-Yet-Unclassified. An in-depth presentation of catalytic sites and mechanisms, based on this classification, is the subject of Chapt. 3. 

Stereoelectronic Factors That Control the Fate of the Tetrahedral Intermediate in Serine Peptidases... 

The previous chapter offered a broad overview of peptidases and esterases in terms of their classification, localization, and some physiological roles. Mention was made of the classification of hydrolases based on a characteristic functionality in their catalytic site, namely serine hydrolases, cysteine hydrolases, aspartic hydrolases, and metallopeptidases. What was left for the present chapter, however, is a detailed presentation of their catalytic site and mechanisms. As such, this chapter serves as a logical link between the preceding overview and the following chapters, whose focus is on metabolic reactions. 

The mechanism by which serine peptidases, particularly serine endopep-tidases (EC 3.4.21), hydrolyze peptide bonds in peptides and proteins has been extensively investigated by X-ray crystallography, site-directed mutagenesis, detection of intermediates, chemical modification, H-NMR spectroscopy, and neutron diffraction [2-14], These studies revealed that all serine peptidases possess a catalytic triad, composed of a serine, a histidine, and an aspartate residue, and a so-called oxyanion hole formed by backbone NH groups. 

It is interesting to note that serine peptidases can, under special conditions in vitro, catalyze the reverse reaction, namely the formation of a peptide bond (Fig. 3.4). The overall mechanism of peptide-bond synthesis by peptidases is represented by the reverse sequence f-a in Fig. 3.3. The nucleophilic amino group of an amino acid residue competes with H20 and reacts with the acyl-enzyme intermediate to form a new peptide bond (Steps d-c in Fig. 3.3). This mechanism is not relevant to the in vivo biosynthesis of proteins but has proved useful for preparative peptide synthesis in vitro [17]. An interesting application of the peptidase-catalyzed peptide synthesis is the enzymatic conversion of porcine insulin to human insulin [18][19]. 

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