Serine protease 636375 219905477 2008-06-17T11:51:41Z 82.17.66.106 [[Image:1UTN.png|thumb|right|250px|[[X-ray crystallography|Crystal structure]] of [[Trypsin]], a typical serine protease.]] In [[biochemistry]], '''serine proteases''' or '''serine endopeptidases''' (newer name) are a class of [[enzyme]]s that cut [[peptide bond]]s in [[protein]]s ([[peptidase]]s). One of the [[amino acids]] at the active site of the enzyme is always a [[serine]], which is where they get their name. They are found in both single-cell and complex organisms, in both cells with nuclei ([[eukaryotes]]) and without nuclei [[prokaryotes]]). Serine proteases are grouped into clans that share structural similarities (homology) and are then further subgrouped into families with simiar sequences. The major clans found in humans include the [[chymotrypsin]]-like, the [[subtilisin]]-like, the alpha/beta hydrolase, and signal peptidase clans. In evolutionary history, serine proteases were originally [[digestion|digestive]] enzymes. In mammals, they evolved by gene duplication to serve functions in [[coagulation|blood clotting]], the [[immunity (medical)|immune system]], and [[inflammation]]. Serine proteases are paired with serine protease inhibitors, which turn off their activity when they are no longer needed.<ref>[http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/S/Serine_Proteases.html#Serpins Kimball's Biology Pages, Serine Proteases]</ref> ==Digestive serine proteases== ===Members=== ====Chymotrypsin-clan==== The three serine proteases of the chymotrypsin-like clan that have been studied in greatest detail are [[chymotrypsin]], [[trypsin]], and [[elastase]]. All three [[enzymes]] are synthesized by the [[pancreas|pancreatic]] acinar cells, secreted in the [[small intestine]] and are responsible for catalyzing the [[hydrolysis]] of [[peptide bond]]s. All three of these enzymes are similar in structure, as shown through their [[X-ray structure]]s. The differing aspect lies in the peptide bond which is being cleaved, which is called the [[scissile]] bond. The different [[enzymes]], like most enzymes, are highly specific in the reactions they catalyze. Each of these digestive serine proteases targets different regions of a [[polypeptide]] chain, based upon the side chains of the amino acid residues surrounding the site of cleavage: *[[Chymotrypsin]] is responsible for cleaving peptide bonds following a ''bulky hydrophobic'' amino acid residue. Preferred residues include [[phenylalanine]], [[tryptophan]] and [[tyrosine]], which fit into a snug [[hydrophobic]] pocket. *[[Trypsin]] is responsible for cleaving peptide bonds following a ''positively-charged'' amino acid residue. Instead of having the [[hydrophobic]] pocket of the ''chymotrypsin'', there exists an [[aspartic acid]] residue at the base of the pocket. This can then interact with positively-charged residues such as [[arginine]] and [[lysine]] on the substrate peptide to be cleaved. *[[Elastase]] is responsible for cleaving peptide bonds following a ''small neutral'' amino acid residue, such as [[Alanine]], [[glycine]] and [[valine]]. (These amino acid residues form much of the connective tissues in meat). The pocket that is in "trypsin" and "chymotrypsin" is now partially filled with [[valine]] and [[threonine]], rendering it a mere depression, which can accommodate these smaller amino acid residues. The combination of these three enzymes make an incredibly effective digestive team, and are primarily responsible for the digestion of [[proteins]]. ====Subtilisin==== Subtilisin is a serine protease in [[prokaryotes]]. Subtilisin is evolutionary unrelated to the chymotrypsin-clan, but shares the same catalytic mechanism utilising a [[catalytic triad]], to create a nucleophilic [[serine]]. This is the classic example used to illustrate [[convergent evolution]], since the same mechanism evolved twice independently during [[evolution]]. ===Catalytic mechanism=== The main player in the catalytic mechanism in the chymotrypsin and subtillisin clan enzymes mentioned above is the ''[[catalytic triad]]''. The triad is located in the active site of the enzyme, where catalysis occurs, and is preserved in all serine protease enzymes. The triad is a coordinated structure consisting of three essential [[amino acids]]: [[histidine]] (His 57), [[serine]] (Ser 195) (hence the name "serine protease") and [[aspartic acid]] (Asp 102). Located very near one another near the heart of the enzyme, these three key amino acids each play an essential role in the cleaving ability of the proteases. In the event of catalysis, an ordered mechanism occurs in which several intermediates are generated. The catalysis of the peptide cleavage can be seen as a [[ping-pong]] catalysis, in which a [[substrate (biochemistry)|substrate]] binds (in this case, the polypeptide being cleaved), a product is released (the N-terminus "half" of the peptide), another substrate binds (in this case, water), and another product is released (the C-terminus "half" of the peptide). Each amino acid in the triad performs a specific task in this process: [[Image:Serine protease mechanism by snellios.png|400px|right|serine protease reaction mechanism]]*The [[serine]] has an -OH group that is able to act as a [[nucleophile]], attacking the [[carbonyl]] carbon of the scissile [[peptide bond]] of the substrate. *A pair of electrons on the [[histidine]] nitrogen has the ability to accept the [[hydrogen]] from the [[serine]] -OH group, thus coordinating the attack of the [[peptide bond]]. *The [[carboxyl]] group on the [[aspartic acid]] in turn [[hydrogen bonds]] with the [[histidine]], making the pair of electrons mentioned above much more [[electronegative]]. The whole reaction can be summarized as follows: *The [[polypeptide]] substrate binds to the surface of the serine protease enzyme such that scissile bond is inserted into the active site of the enzyme, with the carbonyl carbon of this bond positioned near the [[nucleophilic]] [[serine]]. *The [[serine]] -OH attacks the [[carbonyl]] carbon, and the nitrogen of the [[histidine]] accepts the hydrogen from the -OH of the [serine] and a pair of electrons from the double bond of the [[carbonyl]] oxygen moves to the oxygen. As a result, a tetrahedral intermediate is generated. *The bond joining the nitrogen and the carbon in the peptide bond is now broken. The [[covalent electron]]s creating this bond move to attack the hydrogen of the [[histidine]], breaking the connection. The electrons that previously moved from the [[carbonyl]] oxygen double bond move back from the negative oxygen to recreate the bond, generating an [[acyl-enzyme intermediate]]. *Now, water comes in to the reaction. Water replaces the [[N-terminus]] of the cleaved peptide, and attacks the [[carbonyl]] carbon. Once again, the electrons from the double bond move to the oxygen making it negative, as the bond between the oxygen of the water and the carbon is formed. This is coordinated by the nitrogen of the [[histidine]]. which accepts a proton from the water. Overall, this generates another tetrahedral intermediate. *In a final reaction, the bond formed in the first step between the [[serine]] and the [[carbonyl]] carbon moves to attack the hydrogen that the [[histidine]] just acquired. The now electron-deficient [[carbonyl]] carbon re-forms the double bond with the oxygen. As a result, the [[C-terminus]] of the peptide is now ejected. ===Additional stabilizing effects=== It was discovered that additional amino acids of the protease, ''Gly 193'' and ''Ser 195'', are involved in creating what is called an ''[[oxyanion]] hole''. Both ''Gly 193'' and ''Ser 195'' can donate backbone hydrogens for hydrogen bonding. When the [[tetrahedral intermediate]] of step 1 and step 3 are generated, the negative oxygen ion, having accepted the electrons from the [[carbonyl]] double bond fits perfectly into the oxyanion hole. In effect, serine proteases preferentially bind the [[transition state]] and the overall structure is favored, lowering the activation energy of the reaction. This "preferential binding" is responsible for much of the catalytic efficiency of the enzyme. === Zymogens === There are certain [[Enzyme inhibitor|inhibitor]]s which resemble the tetrahedral intermediate, and thus fill up the active site, preventing the enzyme from working properly. Trypsin, a powerful digestive enzyme, is generated in the pancreas. Inhibitors prevent self-digestion of the pancreas itself. [[Zymogen]]s are the usually inactive precursors of an enzyme. If the digestive enzymes were active when synthesized, they would immediately start chewing up the synthesizing organs and tissues. [[Acute pancreatitis]] is such a condition, in which there is premature activation of the digestive enzymes in the pancreas, resulting in self-digestion (autolysis). It also complicates [[postmortem investigation]]s, as the pancreas often digests itself before it can be assessed visually. Zymogens are large, inactive structures, which have the ability to break apart or change into the smaller activated enzymes. The difference between zymogens and the activated enzymes lies in the fact that the active site for catalysis of the zymogens is distorted. As a result, the substrate polypeptide cannot bind effectively, and [[proteolysis]] does not occur. Only after activation, during which the conformation and structure of the zymogen change and the active site is opened, can [[proteolysis]] occur. {| class="wikitable" | '''Zymogen''' || '''Enzyme''' || '''Notes''' |- | ''Trypsinogen'' || [[trypsin]] || When trypsinogen enters the [[small intestine]] from the pancreas, secretions from the [[duodenal mucosa]] cleaves the lysine 15 - isoleucine 16 peptide bond of the zymogen. As a result, the zymogen trypsinogen breaks down into trypsin. Recall that trypsin is also responsible for cleaving [[lysine]] peptide bonds, and thus, once a small amount of trypsin is generated, it participates in cleavage of its own zymogen, generating even more trypsin. The process of trypsin activation can thus be called [[autocatalytic]]. |- | ''Chymotrypsinogen'' || [[chymotrypsin]] || After the Arg 15 - Ile 16 bond in the chymotrypsinogen zymogen is cleaved by trypsin, the newly generated structure called a ''pi-chymotrypsin'' undergoes [[autolysis]] (self digestion), yielding active chymotrypsin. |- | ''Proelastase'' || [[elastase]] || It is activated by cleavage through trypsin. |} As can be seen, trypsinogen activation to ''trypsin'' is essential, because it activates its own reaction, as well as the reaction of both ''chymotrypsin'' and ''elastase''. It is therefore essential that this activation doesn't occur prematurely. There are several protective measures taken by the organism to prevent self-digestion: *The activation of trypsinogen by trypsin is relatively slow *The zymogens are stored in [[zymogen granules]], capsules that have walls that are thought to be resistant to proteolysis. ==Inhibition== Serine proteases are inhibited by a diverse group of [[inhibitor]]s, including synthetic chemical inhibitors for research or therapeutic purposes, and also natural proteinaceous inhibitors. One family of natural inhibitors called "serpins" (abbreviated from [[serine protease inhibitor]]s) can form a [[covalent]] bond with the serine protease, inhibiting its function. The best-studied ''serpins'' are [[antithrombin]] and [[alpha 1-antitrypsin]], studied for their role in [[coagulation]]/[[thrombosis]] and [[emphysema]]/[[A1AT]] respectively. Artificial irreversible small molecule inhibitors include [[AEBSF]] and [[PMSF]]. ==Role in disease== Mutations may lead to decreased or increased activity of enzymes. This may have different consequences, depending on the normal function of the serine protease. For example, mutations in [[protein C]], when leading to insufficient protein levels or activity, predispose to [[thrombosis]]. ==Diagnostic use== Determination of serine protease levels may be useful in the context of particular diseases. * [[Coagulation factor]] levels may be required in the diagnosis of hemorrhagic or thrombotic conditions. * [[Fecal elastase]] is employed to determine the exocrine activity of the pancreas, e.g. in [[cystic fibrosis]] or [[chronic pancreatitis]]. * [[Prostate specific antigen]] is used to determine [[prostate cancer]] risk ==References== <references/> ==External links== * [http://biochem.wustl.edu/~protease/ Serine protease] site of the [[Washington University in St. Louis]] (WUSTL) * {{MeshName|Serine+proteases}} {{Proteases}} {{Serine endopeptidases}} [[Category:EC 3.4.21|*]] [[de:Serinproteinase]] [[it:Serin proteasi]] [[he:סרין פרוטאז]] [[ja:セリンプロテアーゼ]] [[zh:丝氨酸蛋白酶]]