Catalytic triad

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The enzyme TEV protease contains an example of a catalytic triad of residues (red) in its active site. The triad consists of an aspartate (acid), histidine (base) and serine (nucleophile). The substrate (black) is bound by the binding site to orient it next to the triad. (PDB: 1LVM ) Catalytic triad of TEV protease.png
The enzyme TEV protease contains an example of a catalytic triad of residues (red) in its active site. The triad consists of an aspartate (acid), histidine (base) and serine (nucleophile). The substrate (black) is bound by the binding site to orient it next to the triad. ( PDB: 1LVM )

A catalytic triad is a set of three coordinated amino acids that can be found in the active site of some enzymes. [1] [2] Catalytic triads are most commonly found in hydrolase and transferase enzymes (e.g. proteases, amidases, esterases, acylases, lipases and β-lactamases). An acid-base-nucleophile triad is a common motif for generating a nucleophilic residue for covalent catalysis. The residues form a charge-relay network to polarise and activate the nucleophile, which attacks the substrate, forming a covalent intermediate which is then hydrolysed to release the product and regenerate free enzyme. The nucleophile is most commonly a serine or cysteine amino acid, but occasionally threonine or even selenocysteine. The 3D structure of the enzyme brings together the triad residues in a precise orientation, even though they may be far apart in the sequence (primary structure). [3]

Contents

As well as divergent evolution of function (and even the triad's nucleophile), catalytic triads show some of the best examples of convergent evolution. Chemical constraints on catalysis have led to the same catalytic solution independently evolving in at least 23 separate superfamilies. [2] Their mechanism of action is consequently one of the best studied in biochemistry. [4] [5]

History

The enzymes trypsin and chymotrypsin were first purified in the 1930s. [6] A serine in each of trypsin and chymotrypsin was identified as the catalytic nucleophile (by diisopropyl fluorophosphate modification) in the 1950s. [7] The structure of chymotrypsin was solved by X-ray crystallography in the 1960s, showing the orientation of the catalytic triad in the active site. [8] Other proteases were sequenced and aligned to reveal a family of related proteases, [9] [10] [11] now called the S1 family. Simultaneously, the structures of the evolutionarily unrelated papain and subtilisin proteases were found to contain analogous triads. The 'charge-relay' mechanism for the activation of the nucleophile by the other triad members was proposed in the late 1960s. [12] As more protease structures were solved by X-ray crystallography in the 1970s and 80s, homologous (such as TEV protease) and analogous (such as papain) triads were found. [13] [14] [15] The MEROPS classification system in the 1990s and 2000s began classing proteases into structurally related enzyme superfamilies and so acts as a database of the convergent evolution of triads in over 20 superfamilies. [16] [17] Understanding how chemical constraints on evolution led to the convergence of so many enzyme families on the same triad geometries has developed in the 2010s. [2]

Since their initial discovery, there have been increasingly detailed investigations of their exact catalytic mechanism. Of particular contention in the 1990s and 2000s was whether low-barrier hydrogen bonding contributed to catalysis, [18] [19] [20] or whether ordinary hydrogen bonding is sufficient to explain the mechanism. [21] [22] The massive body of work on the charge-relay, covalent catalysis used by catalytic triads has led to the mechanism being the best characterised in all of biochemistry. [4] [5] [21]

Function

Enzymes that contain a catalytic triad use it for one of two reaction types: either to split a substrate (hydrolases) or to transfer one portion of a substrate over to a second substrate (transferases). Triads are an inter-dependent set of residues in the active site of an enzyme and act in concert with other residues (e.g. binding site and oxyanion hole) to achieve nucleophilic catalysis. These triad residues act together to make the nucleophile member highly reactive, generating a covalent intermediate with the substrate that is then resolved to complete catalysis.[ citation needed ]

Mechanism

Catalytic triads perform covalent catalysis using a residue as a nucleophile. The reactivity of the nucleophilic residue is increased by the functional groups of the other triad members. The nucleophile is polarised and oriented by the base, which is itself bound and stabilised by the acid.[ citation needed ]

Catalysis is performed in two stages. First, the activated nucleophile attacks the carbonyl carbon and forces the carbonyl oxygen to accept an electron pair, leading to a tetrahedral intermediate. The build-up of negative charge on this intermediate is typically stabilized by an oxyanion hole within the active site. The intermediate then collapses back to a carbonyl, ejecting the first half of the substrate, but leaving the second half still covalently bound to the enzyme as an acyl-enzyme intermediate. Although general-acid catalysis for breakdown of the First and Second tetrahedral intermediate may occur by the path shown in the diagram, evidence supporting this mechanism with chymotrypsin [23] has been controverted. [24]

The second stage of catalysis is the resolution of the acyl-enzyme intermediate by the attack of a second substrate. If this substrate is water then the result is hydrolysis; if it is an organic molecule then the result is transfer of that molecule onto the first substrate. Attack by this second substrate forms a new tetrahedral intermediate, which resolves by ejecting the enzyme's nucleophile, releasing the second product and regenerating free enzyme. [25]

Triad mechanism.svg
General reaction mechanism of catalysed by a catalytic triad (black): nucleophilic substitution at a carbonyl substrate (red) by a second substrate (blue). First, the enzyme's nucleophile (X) attacks the carbonyl to form a covalently linked acyl-enzyme intermediate. This intermediate is then attacked by the second substrate's nucleophile (X'). If the second nucleophile is the hydroxyl of water, the result is hydrolysis, otherwise the result is group transfer of X'.

Identity of triad members

A catalytic triad charge-relay system as commonly found in proteases. The acid residue (commonly glutamate or aspartate) aligns and polarises the base (usually histidine) which activates the nucleophile (often serine or cysteine, occasionally threonine). The triad reduces the pKa of the nucleophilic residue which then attacks the substrate. An oxyanion hole of positively charged usually backbone amides (occasionally side-chains) stabilise charge build-up on the substrate transition state. Triad chemical mech.png
A catalytic triad charge-relay system as commonly found in proteases. The acid residue (commonly glutamate or aspartate) aligns and polarises the base (usually histidine) which activates the nucleophile (often serine or cysteine, occasionally threonine). The triad reduces the pKa of the nucleophilic residue which then attacks the substrate. An oxyanion hole of positively charged usually backbone amides (occasionally side-chains) stabilise charge build-up on the substrate transition state.

Nucleophile

The side-chain of the nucleophilic residue performs covalent catalysis on the substrate. The lone pair of electrons present on the oxygen or sulfur attacks the electropositive carbonyl carbon. [3] The 20 naturally occurring biological amino acids do not contain any sufficiently nucleophilic functional groups for many difficult catalytic reactions. Embedding the nucleophile in a triad increases its reactivity for efficient catalysis. The most commonly used nucleophiles are the hydroxyl (OH) of serine and the thiol/thiolate ion (SH/S) of cysteine. [2] Alternatively, threonine proteases use the secondary hydroxyl of threonine, however due to steric hindrance of the side chain's extra methyl group such proteases use their N-terminal amide as the base, rather than a separate amino acid. [1] [26]

Use of oxygen or sulfur as the nucleophilic atom causes minor differences in catalysis. Compared to oxygen, sulfur's extra d orbital makes it larger (by 0.4 Å) [27] and softer, allows it to form longer bonds (dC-X and dX-H by 1.3-fold), and gives it a lower pKa (by 5 units). [28] Serine is therefore more dependent than cysteine on optimal orientation of the acid-base triad members to reduce its pKa [28] in order to achieve concerted deprotonation with catalysis. [2] The low pKa of cysteine works to its disadvantage in the resolution of the first tetrahedral intermediate as unproductive reversal of the original nucleophilic attack is the more favourable breakdown product. [2] The triad base is therefore preferentially oriented to protonate the leaving group amide to ensure that it is ejected to leave the enzyme sulfur covalently bound to the substrate N-terminus. Finally, resolution of the acyl-enzyme (to release the substrate C-terminus) requires serine to be re-protonated whereas cysteine can leave as S. Sterically, the sulfur of cysteine also forms longer bonds and has a bulkier van der Waals radius [2] and if mutated to serine can be trapped in unproductive orientations in the active site. [27]

Very rarely, the selenium atom of the uncommon amino acid selenocysteine is used as a nucleophile. [29] The deprotonated Se state is strongly favoured when in a catalytic triad. [29]

Base

Since no natural amino acids are strongly nucleophilic, the base in a catalytic triad polarises and deprotonates the nucleophile to increase its reactivity. [3] Additionally, it protonates the first product to aid leaving group departure.[ citation needed ]

The base is most commonly histidine since its pKa allows for effective base catalysis, hydrogen bonding to the acid residue, and deprotonation of the nucleophile residue. [1] β-lactamases such as TEM-1 use a lysine residue as the base. Because lysine's pKa is so high (pKa=11), a glutamate and several other residues act as the acid to stabilise its deprotonated state during the catalytic cycle. [30] [31] Threonine proteases use their N-terminal amide as the base, since steric crowding by the catalytic threonine's methyl prevents other residues from being close enough. [32] [33]

Acid

The acidic triad member forms a hydrogen bond with the basic residue. This aligns the basic residue by restricting its side-chain rotation, and polarises it by stabilising its positive charge. [3] Two amino acids have acidic side chains at physiological pH (aspartate or glutamate) and so are the most commonly used for this triad member. [3] Cytomegalovirus protease [lower-alpha 2] uses a pair of histidines, one as the base, as usual, and one as the acid. [1] The second histidine is not as effective an acid as the more common aspartate or glutamate, leading to a lower catalytic efficiency. In some enzymes, the acid member of the triad is less necessary and some act only as a dyad. For example, papain [lower-alpha 3] uses asparagine as its third triad member which orients the histidine base but does not act as an acid. Similarly, hepatitis A virus protease [lower-alpha 4] contains an ordered water in the position where an acid residue should be.[ citation needed ]

Examples of triads

The range of amino acid residues used in different combinations in different enzymes to make up a catalytic triad for hydrolysis. On the left are the nucleophile, base and acid triad members. On the right are different substrates with the cleaved bond indicated by a pair of scissors. Two different bonds in beta-lactams can be cleaved (1 by penicillin acylase and 2 by beta-lactamase). Triads and their substrates annotated.svg
The range of amino acid residues used in different combinations in different enzymes to make up a catalytic triad for hydrolysis. On the left are the nucleophile, base and acid triad members. On the right are different substrates with the cleaved bond indicated by a pair of scissors. Two different bonds in beta-lactams can be cleaved (1 by penicillin acylase and 2 by beta-lactamase).

Ser-His-Asp

The Serine-Histidine-Aspartate motif is one of the most thoroughly characterised catalytic motifs in biochemistry. [3] The triad is exemplified by chymotrypsin, [lower-alpha 5] a model serine protease from the PA superfamily which uses its triad to hydrolyse protein backbones. The aspartate is hydrogen bonded to the histidine, increasing the pKa of its imidazole nitrogen from 7 to around 12. This allows the histidine to act as a powerful general base and to activate the serine nucleophile. It also has an oxyanion hole consisting of several backbone amides which stabilises charge build-up on intermediates. The histidine base aids the first leaving group by donating a proton, and also activates the hydrolytic water substrate by abstracting a proton as the remaining OH attacks the acyl-enzyme intermediate.[ citation needed ]

The same triad has also convergently evolved in α/β hydrolases such as some lipases and esterases, however orientation of the triad members is reversed. [34] [35] Additionally, brain acetyl hydrolase (which has the same fold as a small G-protein) has also been found to have this triad. The equivalent Ser-His-Glu triad is used in acetylcholinesterase.[ citation needed ]

Cys-His-Asp

The second most studied triad is the Cysteine-Histidine-Aspartate motif. [2] Several families of cysteine proteases use this triad set, for example TEV protease [lower-alpha 1] and papain. [lower-alpha 3] The triad acts similarly to serine protease triads, with a few notable differences. Due to cysteine's low pKa, the importance of the Asp to catalysis varies and several cysteine proteases are effectively Cys-His dyads (e.g. hepatitis A virus protease), whilst in others the cysteine is already deprotonated before catalysis begins (e.g. papain). [36] This triad is also used by some amidases, such as N-glycanase to hydrolyse non-peptide C-N bonds. [37]

Ser-His-His

The triad of cytomegalovirus protease [lower-alpha 2] uses histidine as both the acid and base triad members. Removing the acid histidine results in only a 10-fold activity loss (compared to >10,000-fold when aspartate is removed from chymotrypsin). This triad has been interpreted as a possible way of generating a less active enzyme to control cleavage rate. [26]

Ser-Glu-Asp

An unusual triad is found in seldolisin proteases. [lower-alpha 6] The low pKa of the glutamate carboxylate group means that it only acts as a base in the triad at very low pH. The triad is hypothesised to be an adaptation to specific environments like acidic hot springs (e.g. kumamolysin) or cell lysosome (e.g. tripeptidyl peptidase). [26]

Cys-His-Ser

The endothelial protease vasohibin [lower-alpha 7] uses a cysteine as the nucleophile, but a serine to coordinate the histidine base. [38] [39] Despite the serine being a poor acid, it is still effective in orienting the histidine in the catalytic triad. [38] Some homologues alternatively have a threonine instead of serine at the acid location. [38]

Thr-Nter, Ser-Nter and Cys-Nter

Threonine proteases, such as the proteasome protease subunit [lower-alpha 8] and ornithine acyltransferases [lower-alpha 9] use the secondary hydroxyl of threonine in a manner analogous to the use of the serine primary hydroxyl. [32] [33] However, due to the steric interference of the extra methyl group of threonine, the base member of the triad is the N-terminal amide which polarises an ordered water which, in turn, deprotonates the catalytic hydroxyl to increase its reactivity. [1] [26] Similarly, there exist equivalent 'serine only' and 'cysteine only' configurations such as penicillin acylase G [lower-alpha 10] and penicillin acylase V [lower-alpha 11] which are evolutionarily related to the proteasome proteases. Again, these use their N-terminal amide as a base. [26]

Ser-cisSer-Lys

This unusual triad occurs only in one superfamily of amidases. In this case, the lysine acts to polarise the middle serine. [40] The middle serine then forms two strong hydrogen bonds to the nucleophilic serine to activate it (one with the side chain hydroxyl and the other with the backbone amide). The middle serine is held in an unusual cis orientation to facilitate precise contacts with the other two triad residues. The triad is further unusual in that the lysine and cis-serine both act as the base in activating the catalytic serine, but the same lysine also performs the role of the acid member as well as making key structural contacts. [40] [41]

Sec-His-Glu

The rare, but naturally occurring amino acid selenocysteine (Sec), can also be found as the nucleophile in some catalytic triads. [29] Selenocysteine is similar to cysteine, but contains a selenium atom instead of a sulfur. An example is in the active site of thioredoxin reductase, which uses the selenium for reduction of disulfide in thioredoxin. [29]

Engineered triads

In addition to naturally occurring types of catalytic triads, protein engineering has been used to create enzyme variants with non-native amino acids, or entirely synthetic amino acids. [42] Catalytic triads have also been inserted into otherwise non-catalytic proteins, or protein mimics.[ citation needed ]

Subtilisin (a serine protease) has had its oxygen nucleophile replaced with each of sulfur, [43] [44] selenium, [45] or tellurium. [46] Cysteine and selenocysteine were inserted by mutagenesis, whereas the non-natural amino acid, tellurocysteine, was inserted using auxotrophic cells fed with synthetic tellurocysteine. These elements are all in the 16th periodic table column (chalcogens), so have similar properties. [47] [48] In each case, changing the nucleophile reduced the enzyme's protease activity, but increased a different activity. A sulfur nucleophile improved the enzymes transferase activity (sometimes called subtiligase). Selenium and tellurium nucleophiles converted the enzyme into an oxidoreductase. [45] [46] When the nucleophile of TEV protease was converted from cysteine to serine, it protease activity was strongly reduced, but was able to be restored by directed evolution. [49]

Non-catalytic proteins have been used as scaffolds, having catalytic triads inserted into them which were then improved by directed evolution. The Ser-His-Asp triad has been inserted into an antibody, [50] as well as a range of other proteins. [51] Similarly, catalytic triad mimics have been created in small organic molecules like diaryl diselenide, [52] [53] and displayed on larger polymers like Merrifield resins, [54] and self-assembling short peptide nanostructures. [55]

Divergent evolution

The sophistication of the active site network causes residues involved in catalysis (and residues in contact with these) to be highly evolutionarily conserved. [56] However, there are examples of divergent evolution in catalytic triads, both in the reaction catalysed, and the residues used in catalysis. The triad remains the core of the active site, but it is evolutionarily adapted to serve different functions. [57] [58] Some proteins, called pseudoenzymes, have non-catalytic functions (e.g. regulation by inhibitory binding) and have accumulated mutations that inactivate their catalytic triad. [59]

Reaction changes

Catalytic triads perform covalent catalysis via an acyl-enzyme intermediate. If this intermediate is resolved by water, the result is hydrolysis of the substrate. However, if the intermediate is resolved by attack by a second substrate, then the enzyme acts as a transferase. For example, attack by an acyl group results in an acyltransferase reaction. Several families of transferase enzymes have evolved from hydrolases by adaptation to exclude water and favour attack of a second substrate. [60] In different members of the α/β-hydrolase superfamily, the Ser-His-Asp triad is tuned by surrounding residues to perform at least 17 different reactions. [35] [61] Some of these reactions are also achieved with mechanisms that have altered formation, or resolution of the acyl-enzyme intermediate, or that don't proceed via an acyl-enzyme intermediate. [35]

Additionally, an alternative transferase mechanism has been evolved by amidophosphoribosyltransferases, which has two active sites. [lower-alpha 12] In the first active site, a cysteine triad hydrolyses a glutamine substrate to release free ammonia. The ammonia then diffuses though an internal tunnel in the enzyme to the second active site, where it is transferred to a second substrate. [62] [63]

Nucleophile changes

Divergent evolution of PA clan proteases to use different nucleophiles in their catalytic triad. Shown are the serine triad of chymotrypsin and the cysteine triad of TEV protease. (PDB: 1LVM, 1GG6 ) Triad divergence.svg
Divergent evolution of PA clan proteases to use different nucleophiles in their catalytic triad. Shown are the serine triad of chymotrypsin and the cysteine triad of TEV protease. ( PDB: 1LVM, 1GG6 )

Divergent evolution of active site residues is slow, due to strong chemical constraints. Nevertheless, some protease superfamilies have evolved from one nucleophile to another. This can be inferred when a superfamily (with the same fold) contains families that use different nucleophiles. [49] Such nucleophile switches have occurred several times during evolutionary history, however the mechanisms by which this happen are still unclear. [17] [49]

Within protease superfamilies that contain a mixture of nucleophiles (e.g. the PA clan), families are designated by their catalytic nucleophile (C=cysteine proteases, S=serine proteases).

Superfamilies containing a mixture of families that use different nucleophiles  [64]
Superfamily Families Examples
PA clan C3, C4, C24, C30, C37, C62, C74, C99TEV protease ( Tobacco etch virus )
S1, S3, S6, S7, S29, S30, S31, S32, S39, S46, S55, S64, S65, S75 Chymotrypsin (mammals, e.g. Bos taurus )
PB clanC44, C45, C59, C69, C89, C95 Amidophosphoribosyltransferase precursor ( Homo sapiens )
S45, S63 Penicillin G acylase precursor ( Escherichia coli )
T1, T2, T3, T6Archaean proteasome, beta component ( Thermoplasma acidophilum )
PC clanC26, C56 Gamma-glutamyl hydrolase ( Rattus norvegicus )
S51 Dipeptidase E ( Escherichia coli )
PD clanC46 Hedgehog protein ( Drosophila melanogaster )
N9, N10, N11 Intein-containing V-type proton ATPase catalytic subunit A ( Saccharomyces cerevisiae )
PE clanP1 DmpA aminopeptidase ( Brucella anthropi )
T5 Ornithine acetyltransferase precursor ( Saccharomyces cerevisiae )

Pseudoenzymes

A further subclass of catalytic triad variants are pseudoenzymes, which have triad mutations that make them catalytically inactive, but able to function as binding or structural proteins. [65] [66] For example, the heparin-binding protein Azurocidin is a member of the PA clan, but with a glycine in place of the nucleophile and a serine in place of the histidine. [67] Similarly, RHBDF1 is a homolog of the S54 family rhomboid proteases with an alanine in the place of the nucleophilic serine. [68] [69] In some cases, pseudoenzymes may still have an intact catalytic triad but mutations in the rest of the protein remove catalytic activity. The CA clan contains catalytically inactive members with mutated triads (calpamodulin has lysine in place of its cysteine nucleophile) and with intact triads but inactivating mutations elsewhere (rat testin retains a Cys-His-Asn triad). [70]

Superfamilies containing pseudoenzymes with inactive triads [65]
Superfamily Families containing pseudoenzymesExamples
CA clan C1, C2, C19 Calpamodulin
CD clanC14 CFLAR
SC clanS9, S33 Neuroligin
SK clanS14 ClpR
SR clanS60Serotransferrin domain 2
ST clanS54 RHBDF1
PA clan S1 Azurocidin 1
PB clanT1 PSMB3

Convergent evolution

Triad convergence ser cys.svg
Evolutionary convergence of serine and cysteine protease towards the same catalytic triads organisation of acid-base-nucleophile in different protease superfamilies. Shown are the triads of subtilisin, [lower-alpha 13] prolyl oligopeptidase, [lower-alpha 14] TEV protease, [lower-alpha 1] and papain. [lower-alpha 3] ( PDB: 1ST2, 1LVM, 3EQ8, 1PE6 )
Triad convergence thr.svg
Evolutionary convergence of threonine proteases towards the same N-terminal active site organisation. Shown are the catalytic threonine of the proteasome [lower-alpha 8] and ornithine acetyltransferase. [lower-alpha 9] ( PDB: 1VRA, 1PMA )

The enzymology of proteases provides some of the clearest known examples of convergent evolution. The same geometric arrangement of triad residues occurs in over 20 separate enzyme superfamilies. Each of these superfamilies is the result of convergent evolution for the same triad arrangement within a different structural fold. This is because there are limited productive ways to arrange three triad residues, the enzyme backbone and the substrate. These examples reflect the intrinsic chemical and physical constraints on enzymes, leading evolution to repeatedly and independently converge on equivalent solutions. [1] [2]

Cysteine and serine hydrolases

The same triad geometries been converged upon by serine proteases such as the chymotrypsin [lower-alpha 5] and subtilisin superfamilies. Similar convergent evolution has occurred with cysteine proteases such as viral C3 protease and papain [lower-alpha 3] superfamilies. These triads have converged to almost the same arrangement due to the mechanistic similarities in cysteine and serine proteolysis mechanisms. [2]

Families of cysteine proteases

Superfamily Families Examples
CA C1, C2, C6, C10, C12, C16, C19, C28, C31, C32, C33, C39, C47, C51, C54, C58, C64, C65, C66, C67, C70, C71, C76, C78, C83, C85, C86, C87, C93, C96, C98, C101 Papain ( Carica papaya ) and calpain ( Homo sapiens )
CDC11, C13, C14, C25, C50, C80, C84 Caspase-1 ( Rattus norvegicus ) and separase ( Saccharomyces cerevisiae )
CEC5, C48, C55, C57, C63, C79 Adenain (human adenovirus type 2)
CFC15 Pyroglutamyl-peptidase I ( Bacillus amyloliquefaciens )
CLC60, C82 Sortase A ( Staphylococcus aureus )
CMC18 Hepatitis C virus peptidase 2 (hepatitis C virus)
CNC9 Sindbis virus-type nsP2 peptidase (sindbis virus)
COC40 Dipeptidyl-peptidase VI ( Lysinibacillus sphaericus )
CPC97 DeSI-1 peptidase ( Mus musculus )
PA C3, C4, C24, C30, C37, C62, C74, C99TEV protease ( Tobacco etch virus )
PBC44, C45, C59, C69, C89, C95 Amidophosphoribosyltransferase precursor ( Homo sapiens )
PCC26, C56 Gamma-glutamyl hydrolase ( Rattus norvegicus )
PDC46 Hedgehog protein ( Drosophila melanogaster )
PEP1 DmpA aminopeptidase ( Brucella anthropi )
unassignedC7, C8, C21, C23, C27, C36, C42, C53, C75

Families of serine proteases

Superfamily Families Examples
SBS8, S53 Subtilisin ( Bacillus licheniformis )
SCS9, S10, S15, S28, S33, S37 Prolyl oligopeptidase ( Sus scrofa )
SES11, S12, S13 D-Ala-D-Ala peptidase C ( Escherichia coli )
SFS24, S26 Signal peptidase I ( Escherichia coli )
SHS21, S73, S77, S78, S80Cytomegalovirus assemblin (human herpesvirus 5)
SJS16, S50, S69 Lon-A peptidase ( Escherichia coli )
SKS14, S41, S49 Clp protease ( Escherichia coli )
SOS74Phage GA-1 neck appendage CIMCD self-cleaving protein (Bacillus phage GA-1)
SPS59 Nucleoporin 145 ( Homo sapiens )
SRS60 Lactoferrin ( Homo sapiens )
SSS66 Murein tetrapeptidase LD-carboxypeptidase ( Pseudomonas aeruginosa )
STS54 Rhomboid-1 ( Drosophila melanogaster )
PA S1, S3, S6, S7, S29, S30, S31, S32, S39, S46, S55, S64, S65, S75 Chymotrypsin A ( Bos taurus )
PBS45, S63 Penicillin G acylase precursor ( Escherichia coli )
PCS51 Dipeptidase E ( Escherichia coli )
PEP1 DmpA aminopeptidase ( Brucella anthropi )
unassignedS48, S62, S68, S71, S72, S79, S81

Threonine proteases

Threonine proteases use the amino acid threonine as their catalytic nucleophile. Unlike cysteine and serine, threonine is a secondary hydroxyl (i.e. has a methyl group). This methyl group greatly restricts the possible orientations of triad and substrate as the methyl clashes with either the enzyme backbone or histidine base. [2] When the nucleophile of a serine protease was mutated to threonine, the methyl occupied a mixture of positions, most of which prevented substrate binding. [71] Consequently, the catalytic residue of a threonine protease is located at its N-terminus. [2]

Two evolutionarily independent enzyme superfamilies with different protein folds are known to use the N-terminal residue as a nucleophile: Superfamily PB (proteasomes using the Ntn fold) [32] and Superfamily PE (acetyltransferases using the DOM fold) [33] This commonality of active site structure in completely different protein folds indicates that the active site evolved convergently in those superfamilies. [2] [26]

Families of threonine proteases

Superfamily Families Examples
PB clanT1, T2, T3, T6Archaean proteasome, beta component ( Thermoplasma acidophilum )
PE clanT5Ornithine acetyltransferase ( Saccharomyces cerevisiae )

See also

Related Research Articles

Chymotrypsin Digestive enzyme

Chymotrypsin (EC 3.4.21.1, chymotrypsins A and B, alpha-chymar ophth, avazyme, chymar, chymotest, enzeon, quimar, quimotrase, alpha-chymar, alpha-chymotrypsin A, alpha-chymotrypsin) is a digestive enzyme component of pancreatic juice acting in the duodenum, where it performs proteolysis, the breakdown of proteins and polypeptides. Chymotrypsin preferentially cleaves peptide amide bonds where the side chain of the amino acid N-terminal to the scissile amide bond (the P1 position) is a large hydrophobic amino acid (tyrosine, tryptophan, and phenylalanine). These amino acids contain an aromatic ring in their side chain that fits into a hydrophobic pocket (the S1 position) of the enzyme. It is activated in the presence of trypsin. The hydrophobic and shape complementarity between the peptide substrate P1 side chain and the enzyme S1 binding cavity accounts for the substrate specificity of this enzyme. Chymotrypsin also hydrolyzes other amide bonds in peptides at slower rates, particularly those containing leucine and methionine at the P1 position.

Trypsin Family of digestive enzymes

Trypsin is an enzyme in the first section of the small intestine that starts the digestion of protein molecules by cutting these long chains of amino acids into smaller pieces. It is a serine protease from the PA clan superfamily, found in the digestive system of many vertebrates, where it hydrolyzes proteins. Trypsin is formed in the small intestine when its proenzyme form, the trypsinogen produced by the pancreas, is activated. Trypsin cuts peptide chains mainly at the carboxyl side of the amino acids lysine or arginine. It is used for numerous biotechnological processes. The process is commonly referred to as trypsin proteolysis or trypsinization, and proteins that have been digested/treated with trypsin are said to have been trypsinized. Trypsin was discovered in 1876 by Wilhelm Kühne and was named from the Ancient Greek word for rubbing since it was first isolated by rubbing the pancreas with glycerin.

Protease Enzyme that cleaves other proteins into smaller peptides

A protease is an enzyme that catalyzes proteolysis, breaking down proteins into smaller polypeptides or single amino acids, and spurring the formation of new protein products. They do this by cleaving the peptide bonds within proteins by hydrolysis, a reaction where water breaks bonds. Proteases are involved in many biological functions, including digestion of ingested proteins, protein catabolism, and cell signaling.

Active site Active region of an enzyme

In biology and biochemistry, the active site is the region of an enzyme where substrate molecules bind and undergo a chemical reaction. The active site consists of amino acid residues that form temporary bonds with the substrate and residues that catalyse a reaction of that substrate. Although the active site occupies only ~10–20% of the volume of an enzyme, it is the most important part as it directly catalyzes the chemical reaction. It usually consists of three to four amino acids, while other amino acids within the protein are required to maintain the tertiary structure of the enzymes.

In biology and biochemistry, protease inhibitors, or antiproteases, are molecules that inhibit the function of proteases. Many naturally occurring protease inhibitors are proteins.

Serine protease Class of enzymes

Serine proteases are enzymes that cleave peptide bonds in proteins. Serine serves as the nucleophilic amino acid at the (enzyme's) active site. They are found ubiquitously in both eukaryotes and prokaryotes. Serine proteases fall into two broad categories based on their structure: chymotrypsin-like (trypsin-like) or subtilisin-like.

A metalloproteinase, or metalloprotease, is any protease enzyme whose catalytic mechanism involves a metal. An example is ADAM12 which plays a significant role in the fusion of muscle cells during embryo development, in a process known as myogenesis.

Cysteine protease Class of enzymes

Cysteine proteases, also known as thiol proteases, are hydrolase enzymes that degrade proteins. These proteases share a common catalytic mechanism that involves a nucleophilic cysteine thiol in a catalytic triad or dyad.

Subtilisin Proteolytic enzyme found in Bacillus subtilis

Subtilisin is a protease initially obtained from Bacillus subtilis.

Enzyme catalysis Catalysis of chemical reactions by specialized proteins known as enzymes

Enzyme catalysis is the increase in the rate of a process by a biological molecule, an "enzyme". Most enzymes are proteins, and most such processes are chemical reactions. Within the enzyme, generally catalysis occurs at a localized site, called the active site.

Aspartic protease

Aspartic proteases are a catalytic type of protease enzymes that use an activated water molecule bound to one or more aspartate residues for catalysis of their peptide substrates. In general, they have two highly conserved aspartates in the active site and are optimally active at acidic pH. Nearly all known aspartyl proteases are inhibited by pepstatin.

Serine hydrolases are one of the largest known enzyme classes comprising approximately ~200 enzymes or 1% of the genes in the human proteome. A defining characteristic of these enzymes is the presence of a particular serine at the active site, which is used for the hydrolysis of substrates. The hydrolysis of the ester or peptide bond proceeds in two steps. First, the acyl part of the substrate is transferred to the serine, making a new ester or amide bond and releasing the other part of the substrate is released. Later, in a slower step, the bond between the serine and the acyl group is hydrolyzed by water or hydroxide ion, regenerating free enzyme. Unlike other, non-catalytic, serines, the reactive serine of these hydrolases is typically activated by a proton relay involving a catalytic triad consisting of the serine, an acidic residue and a basic residue, although variations on this mechanism exist.

TEV protease

TEV protease is a highly sequence-specific cysteine protease from Tobacco Etch Virus (TEV). It is a member of the PA clan of chymotrypsin-like proteases. Due to its high sequence specificity it is frequently used for the controlled cleavage of fusion proteins in vitro and in vivo.

Threonine protease

Threonine proteases are a family of proteolytic enzymes harbouring a threonine (Thr) residue within the active site. The prototype members of this class of enzymes are the catalytic subunits of the proteasome, however the acyltransferases convergently evolved the same active site geometry and mechanism.

3C-like protease Class of enzymes

The 3C-like protease (3CLpro) or main protease (Mpro), formally known as C30 endopeptidase or 3-chymotrypsin-like protease, is the main protease found in coronaviruses. It cleaves the coronavirus polyprotein at eleven conserved sites. It is a cysteine protease and a member of the PA clan of proteases. It has a cysteine-histidine catalytic dyad at its active site and cleaves a Gln–(Ser/Ala/Gly) peptide bond.

A protein superfamily is the largest grouping (clade) of proteins for which common ancestry can be inferred. Usually this common ancestry is inferred from structural alignment and mechanistic similarity, even if no sequence similarity is evident. Sequence homology can then be deduced even if not apparent. Superfamilies typically contain several protein families which show sequence similarity within each family. The term protein clan is commonly used for protease and glycosyl hydrolases superfamilies based on the MEROPS and CAZy classification systems.

PA clan of proteases

The PA clan is the largest group of proteases with common ancestry as identified by structural homology. Members have a chymotrypsin-like fold and similar proteolysis mechanisms but can have identity of <10%. The clan contains both cysteine and serine proteases. PA clan proteases can be found in plants, animals, fungi, eubacteria, archaea and viruses.

Glutamic protease

Glutamic proteases are a group of proteolytic enzymes containing a glutamic acid residue within the active site. This type of protease was first described in 2004 and became the sixth catalytic type of protease. Members of this group of protease had been previously assumed to be an aspartate protease, but structural determination showed it to belong to a novel protease family. The first structure of this group of protease was scytalidoglutamic peptidase, the active site of which contains a catalytic dyad, glutamic acid (E) and glutamine (Q), which give rise to the name eqolisin. This group of proteases are found primarily in pathogenic fungi affecting plant and human.

Asparagine peptide lyase are one of the seven groups in which proteases, also termed proteolytic enzymes, peptidases, or proteinases, are classified according to their catalytic residue. The catalytic mechanism of the asparagine peptide lyases involves an asparagine residue acting as nucleophile to perform a nucleophilic elimination reaction, rather than hydrolysis, to catalyse the breaking of a peptide bond.

Papain-like protease

Papain-like proteases are a large protein family of cysteine protease enzymes that share structural and enzymatic properties with the group's namesake member, papain. They are found in all domains of life. In animals, the group is often known as cysteine cathepsins or, in older literature, lysosomal peptidases. In the MEROPS protease enzyme classification system, papain-like proteases form Clan CA. Papain-like proteases share a common catalytic dyad active site featuring a cysteine amino acid residue that acts as a nucleophile.

References

Notes

  1. 1 2 3 4 TEV protease MEROPS: clan PA, family C4
  2. 1 2 Cytomegalovirus protease MEROPS: clan SH, family S21
  3. 1 2 3 4 Papain MEROPS: clan CA, family C1
  4. Hepatitis A virus protease MEROPS: clan PA, family C3
  5. 1 2 3 Chymotrypsin MEROPS: clan PA, family S1
  6. Seldolisin protease MEROPS: clan SB, family 53
  7. Vasohibin protease MEROPS: clan CA
  8. 1 2 Proteasome MEROPS: clan PB, family T1
  9. 1 2 Ornithine acyltransferases MEROPS: clan PE, family T5
  10. Penicillin acylase G MEROPS: clan PB, family S45
  11. Penicillin acylase V MEROPS: clan PB, family C59
  12. amidophosphoribosyltransferase MEROPS: clan PB, family C44
  13. Subtilisin MEROPS: clan SB, family S8
  14. Prolyl oligopeptidase MEROPS: clan SC, family S9

Citations

  1. 1 2 3 4 5 6 Dodson G, Wlodawer A (1998). "Catalytic triads and their relatives". Trends Biochem. Sci. 23 (9): 347–52. doi:10.1016/S0968-0004(98)01254-7. PMID   9787641.
  2. 1 2 3 4 5 6 7 8 9 10 11 12 13 Buller AR, Townsend CA (2013). "Intrinsic evolutionary constraints on protease structure, enzyme acylation, and the identity of the catalytic triad". Proc. Natl. Acad. Sci. U.S.A. 110 (8): E653–61. Bibcode:2013PNAS..110E.653B. doi: 10.1073/pnas.1221050110 . PMC   3581919 . PMID   23382230.
  3. 1 2 3 4 5 6 Stryer L, Berg JM, Tymoczko JL (2002). "9 Catalytic Strategies". Biochemistry (5th ed.). San Francisco: W.H. Freeman. ISBN   9780716749554.
  4. 1 2 Perutz M (1992). Protein structure. New approaches to disease and therapy. New York: W.H. Freeman and Co. ISBN   9780716770213.
  5. 1 2 Neurath H (1994). "Proteolytic enzymes past and present: the second golden era. Recollections, special section in honor of Max Perutz". Protein Sci. 3 (10): 1734–9. doi:10.1002/pro.5560031013. PMC   2142620 . PMID   7849591.
  6. Ohman KP, Hoffman A, Keiser HR (1990). "Endothelin-induced vasoconstriction and release of atrial natriuretic peptides in the rat". Acta Physiol. Scand. 138 (4): 549–56. doi:10.1111/j.1748-1716.1990.tb08883.x. PMID   2141214.
  7. Dixon GH, Kauffman DL, Neurath H (1958). "Amino Acid Sequence in the Region of Diisopropyl Phosphoryl Binding in Dip-Trypsin". J. Am. Chem. Soc. 80 (5): 1260–1. doi:10.1021/ja01538a059.
  8. Matthews BW, Sigler PB, Henderson R, et al. (1967). "Three-dimensional structure of tosyl-α-chymotrypsin". Nature . 214 (5089): 652–656. Bibcode:1967Natur.214..652M. doi:10.1038/214652a0. PMID   6049071. S2CID   4273406.
  9. Walsh KA, Neurath H (1964). "Trypsinogen and Chymotrypsinogen as Homologous Proteins". Proc. Natl. Acad. Sci. U.S.A. 52 (4): 884–9. Bibcode:1964PNAS...52..884W. doi: 10.1073/pnas.52.4.884 . PMC   300366 . PMID   14224394.
  10. de Haën C, Neurath H, Teller DC (1975). "The phylogeny of trypsin-related serine proteases and their zymogens. New methods for the investigation of distant evolutionary relationships". J. Mol. Biol. 92 (2): 225–59. doi:10.1016/0022-2836(75)90225-9. PMID   1142424.
  11. Lesk AM, Fordham WD (1996). "Conservation and variability in the structures of serine proteinases of the chymotrypsin family". J. Mol. Biol. 258 (3): 501–37. doi:10.1006/jmbi.1996.0264. PMID   8642605.
  12. Blow DM, Birktoft JJ, Hartley BS (1969). "Role of a buried acid group in the mechanism of action of chymotrypsin". Nature . 221 (5178): 337–40. Bibcode:1969Natur.221..337B. doi:10.1038/221337a0. PMID   5764436. S2CID   4214520.
  13. Gorbalenya AE, Blinov VM, Donchenko AP (1986). "Poliovirus-encoded proteinase 3C: a possible evolutionary link between cellular serine and cysteine proteinase families". FEBS Lett. 194 (2): 253–7. doi:10.1016/0014-5793(86)80095-3. PMID   3000829. S2CID   23268152.
  14. Bazan JF, Fletterick RJ (1988). "Viral cysteine proteases are homologous to the trypsin-like family of serine proteases: structural and functional implications". Proc. Natl. Acad. Sci. U.S.A. 85 (21): 7872–6. Bibcode:1988PNAS...85.7872B. doi: 10.1073/pnas.85.21.7872 . PMC   282299 . PMID   3186696.
  15. Phan J, Zdanov A, Evdokimov AG, et al. (2002). "Structural basis for the substrate specificity of tobacco etch virus protease". J. Biol. Chem. 277 (52): 50564–72. doi: 10.1074/jbc.M207224200 . PMID   12377789.
  16. Rawlings ND, Barrett AJ (1993). "Evolutionary families of peptidases". Biochem. J. 290 (1): 205–18. doi:10.1042/bj2900205. PMC   1132403 . PMID   8439290.
  17. 1 2 Rawlings ND, Barrett AJ, Bateman A (2010). "MEROPS: the peptidase database". Nucleic Acids Res. 38 (supl_1): D227–33. doi:10.1093/nar/gkp971. PMC   2808883 . PMID   19892822.
  18. Frey PA, Whitt SA, Tobin JB (1994). "A low-barrier hydrogen bond in the catalytic triad of serine proteases". Science . 264 (5167): 1927–30. Bibcode:1994Sci...264.1927F. doi:10.1126/science.7661899. PMID   7661899.
  19. Ash EL, Sudmeier JL, De Fabo EC, et al. (1997). "A low-barrier hydrogen bond in the catalytic triad of serine proteases? Theory versus experiment". Science . 278 (5340): 1128–32. Bibcode:1997Sci...278.1128A. doi:10.1126/science.278.5340.1128. PMID   9353195.
  20. Agback P, Agback T (2018). "Direct evidence of a low barrier hydrogen bond in the catalytic triad of a Serine protease". Sci. Rep. 8 (1): 10078. Bibcode:2018NatSR...810078A. doi:10.1038/s41598-018-28441-7. PMC   6031666 . PMID   29973622.
  21. 1 2 Schutz CN, Warshel A (2004). "The low barrier hydrogen bond (LBHB) proposal revisited: the case of the Asp... His pair in serine proteases". Proteins . 55 (3): 711–23. doi:10.1002/prot.20096. PMID   15103633. S2CID   34229297.
  22. Warshel A, Papazyan A (1996). "Energy considerations show that low-barrier hydrogen bonds do not offer a catalytic advantage over ordinary hydrogen bonds". Proc. Natl. Acad. Sci. U.S.A. 93 (24): 13665–70. Bibcode:1996PNAS...9313665W. doi: 10.1073/pnas.93.24.13665 . PMC   19385 . PMID   8942991.
  23. Fersht, A.R.; Requena, Y (1971). "Mechanism of the Chymotrypsin-Catalyzed Hydrolysis of Amides. pH Dependence of kc and Km.' Kinetic Detection of an Intermediate". J. Am. Chem. Soc. 93 (25): 7079–87. doi:10.1021/ja00754a066. PMID   5133099.
  24. Zeeberg, B; Caswell, M; Caplow, M (1973). "Concerning a reported change in rate-determining step in chymotrypsin catalysis". J. Am. Chem. Soc. 95 (8): 2734–5. doi:10.1021/ja00789a081. PMID   4694533.
  25. Shafee T (2014). Evolvability of a viral protease: experimental evolution of catalysis, robustness and specificity (PhD thesis). University of Cambridge. doi:10.17863/CAM.16528.
  26. 1 2 3 4 5 6 Ekici OD, Paetzel M, Dalbey RE (2008). "Unconventional serine proteases: variations on the catalytic Ser/His/Asp triad configuration". Protein Sci. 17 (12): 2023–37. doi:10.1110/ps.035436.108. PMC   2590910 . PMID   18824507.
  27. 1 2 McGrath ME, Wilke ME, Higaki JN, et al. (1989). "Crystal structures of two engineered thiol trypsins". Biochemistry . 28 (24): 9264–70. doi:10.1021/bi00450a005. PMID   2611228.
  28. 1 2 Polgár L, Asbóth B (1986). "The basic difference in catalyses by serine and cysteine proteinases resides in charge stabilization in the transition state". J. Theor. Biol. 121 (3): 323–6. Bibcode:1986JThBi.121..323P. doi:10.1016/s0022-5193(86)80111-4. PMID   3540454.
  29. 1 2 3 4 Brandt W, Wessjohann LA (2005). "The functional role of selenocysteine (Sec) in the catalysis mechanism of large thioredoxin reductases: proposition of a swapping catalytic triad including a Sec-His-Glu state". ChemBioChem . 6 (2): 386–94. doi:10.1002/cbic.200400276. PMID   15651042. S2CID   25575160.
  30. Damblon C, Raquet X, Lian LY, et al. (1996). "The catalytic mechanism of beta-lactamases: NMR titration of an active-site lysine residue of the TEM-1 enzyme". Proc. Natl. Acad. Sci. U.S.A. 93 (5): 1747–52. Bibcode:1996PNAS...93.1747D. doi: 10.1073/pnas.93.5.1747 . PMC   39852 . PMID   8700829.
  31. Jelsch C, Lenfant F, Masson JM, et al. (1992). "Beta-lactamase TEM1 of E. coli. Crystal structure determination at 2.5 A resolution". FEBS Lett. 299 (2): 135–42. doi: 10.1016/0014-5793(92)80232-6 . PMID   1544485.
  32. 1 2 3 Brannigan JA, Dodson G, Duggleby HJ, et al. (1995). "A protein catalytic framework with an N-terminal nucleophile is capable of self-activation". Nature . 378 (6555): 416–9. Bibcode:1995Natur.378..416B. doi:10.1038/378416a0. PMID   7477383. S2CID   4277904.
  33. 1 2 3 Cheng H, Grishin NV (2005). "DOM-fold: a structure with crossing loops found in DmpA, ornithine acetyltransferase, and molybdenum cofactor-binding domain". Protein Sci. 14 (7): 1902–10. doi:10.1110/ps.051364905. PMC   2253344 . PMID   15937278.
  34. Sun Y, Yin S, Feng Y, et al. (2014). "Molecular basis of the general base catalysis of an α/β-hydrolase catalytic triad". J. Biol. Chem. 289 (22): 15867–79. doi: 10.1074/jbc.m113.535641 . PMC   4140940 . PMID   24737327.
  35. 1 2 3 Rauwerdink A, Kazlauskas RJ (2015). "How the Same Core Catalytic Machinery Catalyzes 17 Different Reactions: the Serine-Histidine-Aspartate Catalytic Triad of α/β-Hydrolase Fold Enzymes". ACS Catal. 5 (10): 6153–6176. doi:10.1021/acscatal.5b01539. PMC   5455348 . PMID   28580193.
  36. Beveridge AJ (1996). "A theoretical study of the active sites of papain and S195C rat trypsin: implications for the low reactivity of mutant serine proteinases". Protein Sci. 5 (7): 1355–65. doi:10.1002/pro.5560050714. PMC   2143470 . PMID   8819168.
  37. Allen MD, Buchberger A, Bycroft M (2006). "The PUB domain functions as a p97 binding module in human peptide N-glycanase". J. Biol. Chem. 281 (35): 25502–8. doi: 10.1074/jbc.M601173200 . PMID   16807242.
  38. 1 2 3 Sanchez-Pulido L, Ponting CP (2016). "Vasohibins: new transglutaminase-like cysteine proteases possessing a non-canonical Cys-His-Ser catalytic triad". Bioinformatics . 32 (10): 1441–5. doi:10.1093/bioinformatics/btv761. PMC   4866520 . PMID   26794318.
  39. Sato Y, Sonoda H (2007). "The vasohibin family: a negative regulatory system of angiogenesis genetically programmed in endothelial cells". Arterioscler. Thromb. Vasc. Biol. 27 (1): 37–41. doi: 10.1161/01.atv.0000252062.48280.61 . PMID   17095714.
  40. 1 2 Shin S, Yun YS, Koo HM, et al. (2003). "Characterization of a novel Ser-cisSer-Lys catalytic triad in comparison with the classical Ser-His-Asp triad". J. Biol. Chem. 278 (27): 24937–43. doi: 10.1074/jbc.M302156200 . PMID   12711609.
  41. Cerqueira NM, Moorthy H, Fernandes PA, et al. (2017). "The mechanism of the Ser-(cis)Ser-Lys catalytic triad of peptide amidases". Phys. Chem. Chem. Phys. 19 (19): 12343–12354. Bibcode:2017PCCP...1912343C. doi:10.1039/C7CP00277G. PMID   28453015. Archived from the original on 2017-12-24. Retrieved 2017-12-24.
  42. Toscano MD, Woycechowsky KJ, Hilvert D (2007). "Minimalist active-site redesign: teaching old enzymes new tricks". Angew. Chem. 46 (18): 3212–36. doi:10.1002/anie.200604205. PMID   17450624.
  43. Abrahmsén L, Tom J, Burnier J, et al. (1991). "Engineering subtilisin and its substrates for efficient ligation of peptide bonds in aqueous solution". Biochemistry . 30 (17): 4151–9. CiteSeerX   10.1.1.461.9606 . doi:10.1021/bi00231a007. PMID   2021606.
  44. Jackson DY, Burnier J, Quan C, et al. (1994). "A designed peptide ligase for total synthesis of ribonuclease A with unnatural catalytic residues". Science . 266 (5183): 243–7. Bibcode:1994Sci...266..243J. doi:10.1126/science.7939659. JSTOR   2884761. PMID   7939659.
  45. 1 2 Syed R, Wu ZP, Hogle JM, et al. (1993). "Crystal structure of selenosubtilisin at 2.0-A resolution". Biochemistry . 32 (24): 6157–64. doi:10.1021/bi00075a007. PMID   8512925.
  46. 1 2 Mao S, Dong Z, Liu J, et al. (2005). "Semisynthetic tellurosubtilisin with glutathione peroxidase activity". J. Am. Chem. Soc. 127 (33): 11588–9. doi:10.1021/ja052451v. PMID   16104720.
  47. Devillanova FA, Du Mont W (2013). Handbook of Chalcogen Chemistry. Vol. 1: new perspectives in sulfur, selenium and tellurium (2nd ed.). Cambridge: RSC. ISBN   9781849736237. OCLC   868953797.
  48. Bouroushian M (2010). "Electrochemistry of the Chalcogens". Electrochemistry of Metal Chalcogenides. Monographs in Electrochemistry. Berlin, Heidelberg: Springer. pp. 57–75. doi:10.1007/978-3-642-03967-6_2. ISBN   9783642039669.
  49. 1 2 3 Shafee T, Gatti-Lafranconi P, Minter R, et al. (2015). "Handicap-Recover Evolution Leads to a Chemically Versatile, Nucleophile-Permissive Protease". ChemBioChem . 16 (13): 1866–9. doi:10.1002/cbic.201500295. PMC   4576821 . PMID   26097079.
  50. Okochi N, Kato-Murai M, Kadonosono T, et al. (2007). "Design of a serine protease-like catalytic triad on an antibody light chain displayed on the yeast cell surface". Appl. Microbiol. Biotechnol. 77 (3): 597–603. doi:10.1007/s00253-007-1197-0. PMID   17899065. S2CID   35590920.
  51. Rajagopalan S, Wang C, Yu K, et al. (2014). "Design of activated serine-containing catalytic triads with atomic-level accuracy". Nat. Chem. Biol. 10 (5): 386–91. doi:10.1038/nchembio.1498. PMC   4048123 . PMID   24705591.
  52. Bhowmick D, Mugesh G (2015). "Introduction of a catalytic triad increases the glutathione peroxidase-like activity of diaryl diselenides". Org. Biomol. Chem. 13 (34): 9072–82. doi:10.1039/C5OB01294E. PMID   26220806.
  53. Bhowmick D, Mugesh G (2015). "Insights into the catalytic mechanism of synthetic glutathione peroxidase mimetics". Org. Biomol. Chem. 13 (41): 10262–72. doi:10.1039/c5ob01665g. PMID   26372527.
  54. Nothling MD, Ganesan A, Condic-Jurkic K, et al. (2017). "Simple Design of an Enzyme-Inspired Supported Catalyst Based on a Catalytic Triad". Chem. 2 (5): 732–745. doi: 10.1016/j.chempr.2017.04.004 .
  55. Gulseren G, Khalily MA, Tekinay AB, et al. (2016). "Catalytic supramolecular self-assembled peptide nanostructures for ester hydrolysis". J. Mater. Chem. B . 4 (26): 4605–4611. doi:10.1039/c6tb00795c. hdl: 11693/36666 . PMID   32263403.
  56. Halabi N, Rivoire O, Leibler S, et al. (2009). "Protein sectors: evolutionary units of three-dimensional structure". Cell . 138 (4): 774–86. doi:10.1016/j.cell.2009.07.038. PMC   3210731 . PMID   19703402.
  57. Murzin AG (1998). "How far divergent evolution goes in proteins". Current Opinion in Structural Biology . 8 (3): 380–387. doi:10.1016/S0959-440X(98)80073-0. PMID   9666335.
  58. Gerlt JA, Babbitt PC (2001). "Divergent evolution of enzymatic function: mechanistically diverse superfamilies and functionally distinct suprafamilies". Annu. Rev. Biochem. 70 (1): 209–46. doi:10.1146/annurev.biochem.70.1.209. PMID   11395407.
  59. Murphy JM, Farhan H, Eyers PA (2017). "Bio-Zombie: the rise of pseudoenzymes in biology". Biochem. Soc. Trans. 45 (2): 537–544. doi:10.1042/bst20160400. PMID   28408493.
  60. Stehle F, Brandt W, Stubbs MT, et al. (2009). "Sinapoyltransferases in the light of molecular evolution". Phytochemistry . Evolution of Metabolic Diversity. 70 (15–16): 1652–62. doi:10.1016/j.phytochem.2009.07.023. PMID   19695650.
  61. Dimitriou PS, Denesyuk A, Takahashi S, et al. (2017). "Alpha/beta-hydrolases: A unique structural motif coordinates catalytic acid residue in 40 protein fold families". Proteins . 85 (10): 1845–1855. doi:10.1002/prot.25338. PMID   28643343. S2CID   25363240.
  62. Smith JL (1998). "Glutamine PRPP amidotransferase: snapshots of an enzyme in action". Current Opinion in Structural Biology . 8 (6): 686–94. doi:10.1016/s0959-440x(98)80087-0. PMID   9914248.
  63. Smith JL, Zaluzec EJ, Wery JP, et al. (1994). "Structure of the allosteric regulatory enzyme of purine biosynthesis". Science . 264 (5164): 1427–33. Bibcode:1994Sci...264.1427S. doi:10.1126/science.8197456. PMID   8197456.
  64. "Clans of Mixed (C, S, T) Catalytic Type". www.ebi.ac.uk. MEROPS. Retrieved 20 Dec 2018.
  65. 1 2 Fischer K, Reynolds SL (2015). "Pseudoproteases: mechanisms and function". Biochem. J. 468 (1): 17–24. doi:10.1042/BJ20141506. PMID   25940733.
  66. Todd AE, Orengo CA, Thornton JM (2002). "Sequence and structural differences between enzyme and nonenzyme homologs". Structure . 10 (10): 1435–51. doi:10.1016/s0969-2126(02)00861-4. PMID   12377129.
  67. Iversen LF, Kastrup JS, Bjørn SE, et al. (1997). "Structure of HBP, a multifunctional protein with a serine proteinase fold". Nat. Struct. Biol. 4 (4): 265–8. doi:10.1038/nsb0497-265. PMID   9095193. S2CID   19949043.
  68. Zettl M, Adrain C, Strisovsky K, et al. (2011). "Rhomboid family pseudoproteases use the ER quality control machinery to regulate intercellular signaling". Cell . 145 (1): 79–91. doi:10.1016/j.cell.2011.02.047. PMC   3149277 . PMID   21439629.
  69. Lemberg MK, Adrain C (2016). "Inactive rhomboid proteins: New mechanisms with implications in health and disease". Semin. Cell Dev. Biol. 60: 29–37. doi:10.1016/j.semcdb.2016.06.022. hdl: 10400.7/759 . PMID   27378062.
  70. Cheng CY, Morris I, Bardin CW (1993). "Testins Are Structurally Related to the Mouse Cysteine Proteinase Precursor But Devoid of Any Protease/Anti-Protease Activity". Biochem. Biophys. Res. Commun. 191 (1): 224–231. doi:10.1006/bbrc.1993.1206. PMID   8447824.
  71. Pelc LA, Chen Z, Gohara DW, et al. (2015). "Why Ser and not Thr brokers catalysis in the trypsin fold". Biochemistry . 54 (7): 1457–64. doi:10.1021/acs.biochem.5b00014. PMC   4342846 . PMID   25664608.