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| - | {{Template: Oberholser Sandbox Reservation}} | + | {{Template:Oberholser_Sandbox_Reservation}} |
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| | + | <Structure load='1AKE' size='500' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /> |
| | + | =Description= |
| | + | Adenylate kinase, commonly known as ADK, is a phosphotransferase enzyme. It has two important roles in various organisms. First, ADK plays an important role in nucleotide metabolism and synthesis. It also has a role in cellular energetics and homeostasis by phosphotransfer networks. Adenylate kinase catalyzes the reaction that forms ADP. The reaction is ATP + AMP = 2 ADP. In this catalyzed reaction, ADK molecules bind to AMP molecules and increase its binding affinity for ATP over other phosphate groups. However, adenylate kinase is also found in other molecules such as bacteria and yeast. ADK plays similar roles in bacteria and yeast, in that it involves cellular metabolism and energy. The following images highlight the structure of Adenylate kinase from ''Yersinia pestis'', commonly known as yeast. |
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| | + | =Structure= |
| | + | ADK is a protein made up of 214 amino acids. The <scene name='Sandbox_39/Adenylate_kinase_main_chain2/1'>mainchain</scene> of adenylate kinase can be seen here in purple with the ligand in the middle of the protein. The <scene name='Sandbox_39/Adenylate_kinase_beta_sheets2/1'>secondary structure</scene> contains 12 alpha helices (light blue) and 7 beta sheets (yellow). The <scene name='Sandbox_39/Adenylate_kinase_hbonds2/1'>backbone hydrogen bonds</scene> hold the protein together and give it shape. The <scene name='Sandbox_39/Adenylate_kinase_hydro_resid4/1'>hydrophobic residues</scene> are located towards the center of the protein, away from water. These residues are responsible for the overall conformation of the protein because of their "water fearing" nature. The <scene name='Sandbox_39/Adenylate_kinase_philic_resid2/1'>hydrophilic residues</scene> are located mainly on the surface or outside of the protein where they will have contact with water and other molecules. The hydrophilic residues that are inside the enzyme are necessary for the enzyme to open its active site and allow substrate in. The <scene name='Sandbox_39/Adenylate_kinase_ligand/1'>ligand</scene> is located in the center of protein. The ligand has six <scene name='Sandbox_39/Adenylate_kinase_cat_residues/1'>catalytic residues</scene> that allow it to bind to ADK's target protein (catalytic residues shown in white within the green ligand). These catalytic residues are known as the active site. The catalytic residues consist of arginine, aspartic acid, and lysine. The <scene name='Sandbox_39/Adenylate_kinase_solv_w_lig/1'>ligand has minimal water contact</scene> because it is located in the center of the protein (ligand is orange and water is displayed by space filling model). It is important to be shielded from water so that nothing interferes with the active site of the protein. The waters that are found in the interior are a result of the hydrophobic residues that are also found there. The ligands and catalytic residues of adenylate kinase are highly conserved throughout various organisms. This conservation indicates their importance to the protein's enzymatic function. |
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| - | =='''Papain'''== | + | =References= |
| | + | http://www.pnas.org/content/102/2/303.full |
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| - | <Structure load='9pap' size='300' frame='true' align='right' caption='ribbon structure of papain with methanol solvent molecules' scene=''/>
| + | http://www.uniprot.org/uniprot/O69172 |
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| - | == Introduction ==
| + | http://www.biomedcentral.com/1471-2199/13/31/abstract |
| - | Papain (9PAP), also known as papaya proteinase I, is an enzyme found in unripe papaya fruit. A cysteine protease, it has been used to break down tough muscle fibers, and hence is often found in powdered meat tenderizers. It has also been used in cell isolation procedures because it is very efficient and not very destructive. It is collected from the fruit by scoring its skin and allowing the "sap" to seem out. The sap is then dried and purified.
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| - | The 3D model of papain shown to the right is "decorated" with solvent methanol molecules. The following Jmol representations of papain will not show these solvent molecules.
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| - | == Function ==
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| - | As a cysteine protease, Papain utilizes a nucleophilic cysteine thiol as part of its catalytic triad. Papain's Cys-25 is deprotonated by its His-159. The now nucleophilic Cys-25 attacks the carbonyl carbon of the peptide backbone, forming an acyl enzyme intermediate in which the peptide's amino terminal is free. Also in this step, His-159 is returned to its deprotonated form. The intermediate is then deacylated by a water molecule, and it releases the carboxyl terminal of the peptide to produce the product and regenerate the active enzyme. This entire mechanism is shown below:
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| - | [[Image:jrip.jpg]]
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| - | <ref>Image from:
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| - | http://upload.wikimedia.org/wikipedia/commons/5/5c/Cysteinprotease_Reaktionsmechanismus.svg</ref>]]
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| - | Papain digests most proteins, often more extensively than pancreatic proteases. It has a very broad specificity and is known to cleave peptide bonds of basic amino acids and leucine and glycine residues, but prefers amino acids with large hydrophobic side chains.
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| - | Papain is also known to cleave antibodies above and below the disulfide bonds that join the heavy chains and that is found between the light chain and heavy chain. This generates two monovalent Fab segments, that each have a single antibody binding sites, and an intact Fc fragment, as shown below:
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| - | [[Image:2fab_fc.png|center]]
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| - | <ref>Image from:
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| - | http://en.wikipedia.org/wiki/Papain</ref>]]
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| - | ----
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| - | <Structure load='9pap' size='300' frame='true' align='left' caption='Ribbon Structure of Papain' scene=/>
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| - | | + | |
| - | == Composition of Papain ==
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| - | Proteins only consist of certain elements: carbon, hydrogen, nitrogen, oxygen, and sulfur. Enzymes' primary structures allow them to fold optimally and interact with their substrates maximally in order to efficiently catalyze biological reactions. The <scene name='Sandbox_39/Elemental/2'>elemental composition</scene> of papain, seen to the right as a space-fill model, shows carbon atoms outlined in grey, oxygen atoms in red, nitrogen atoms in blue, and sulfur atoms in yellow.
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| - | In addition, the entirety of the secondary structure of papain can be traced from the <scene name='Sandbox_39/Elemental/3'>N- to C-terminus.</scene> As shown to the left, the red end begins the protein at the
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| - | N-terminus, and can be traced through the colors of the rainbow to the blue end at the C-terminus.
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| - | | + | |
| - | == Primary, Secondary, and Tertiary Structure ==
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| - | Papain is composed of 212 amino acid residues that make up its primary structure. This structure is shown below.
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| - | Papain's primary structure causes it to fold into different motifs that make up its secondary structure. These motifs include <scene name='Sandbox_39/Alpha_helices/2'>alpha helices</scene>, shown in green, and <scene name='Sandbox_39/Beta_pleated_sheets/2'>beta pleated sheets</scene>, shown in orange. As shown to the left, papain has 7 alpha helices and 8 beta pleated sheets. All other motifs are nonrandom, structural units, mostly simply turns.
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| - | Papain's seconary structures then fold even further to for its three-dimensional structure, which consists of two distinct structural domains with a cleft between them. This cleft contains the catalytic diad discussed above. This tertiary structure is pictured to the right.
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| - | [[Image:9pap moreau 1.jpeg|right]]
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| - | <ref>Image from:
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| - | http://linus.chem.ku.edu/hewlett/Chem188/Enzyme/enzyme_background.htm</ref>]]
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| - | == Active Site ==
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| - | Papain has a broad specificity for protein substrates. The active site consists of seven subsites (S1-S4 and S1’-S3’) that can each accommodate one amino acid residue of a protein substrate (P1-P4 and P1’-P3’).
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| - | [[Image:Subsites.jpg|right]]
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| - | <ref>Image from:
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| - | Hans-Hartwig Otto, and Tanja Schirmeister (1997) Cysteine Proteases and Their Inhibitors. Chemical Reviews. No. 97, 133-171.</ref>]]
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| - | Specificity is controlled, however, by the <scene name='Sandbox_39/Active_site_revised/1'>catalytic diad</scene>, a hydrophobic pocket that accommodates the side chains of the protein substrate. This diad, shown above in red and clearly visible in the cleft of the enzyme, consists of cysteine25 (after which the protein is categorized as a cysteine protease)and histidine159. Papain exhibits specific substrate preferences for hydrophobic or aromatic residues.
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| - | == Polarity and Hydrophobicity ==
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| - | Papain contains many hydrophobic, or "water-hating" regions, and hydrophillic, or "water-loving" regions. The hydrophobic effect, or the tendency of nonpolar substances to aggregate in aqueous solution and exclude water molecules, allows proteins to fold the way they do, exposing hydrophilic residues on their outer surface while sequestering hydrophobic residues in their center. As shown above, all of the <scene name='Sandbox_39/Hydrophobic_residues_revised/1'>hydrophobic residues</scene> are found closer to the center of the folded protein, shown at the left in pink, while the <scene name='Sandbox_39/Hydrophillic_residues_revised/1'>hydrophilic residues</scene> are found around the outside of the protein, shown in blue. These spacefill models give a better idea of the surface area of the protein that contacts the aqueous solution.
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| - | == Disulfide Bonds ==
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| - | Papain contains three <scene name='Sandbox_39/Disulfide_bonds/5'>disulfide bonds</scene>. These bonds are found between Cys-22 and Cys 63, between Cys-56 and Cys-95, and between Cys-153 and Cys-200. These bonds can be seen above as yellow rods, connecting Cysteine residues, also shown in yellow.
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| - | == Hydrogen Bonding ==
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| - | Hydrogen bonds are essential for the stability of a protein. In the image shown below, the hydrogen bonds can be seen between the secondary structures of the protein as white, dashed lines. Papain contains very specific hydrogen bonding between the amino acid residues. This representation clearly shows how crucial hydrogen bonding is to help maintain the stability of the protein.
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| - | [[Image:Hbonds.jpg]]
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| - | <ref>Image from:
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| - | http://www.rcsb.org/pdb/explore/jmol.do?structureId=9PAP&bionumber=1</ref>]] | + | |
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| - | == Papain Inhibition ==
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| - | <Structure load='9pap' size='300' frame='true' align='right' caption='Papain/ZLFG-DAM covalent complex' scene=''/>
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| - | Substances that inhibit enzymes have sequences that resemble the normal substrate of that enzyme. Some substances that act to inhibit the enzymatic activity of papain are able to do so because of their structural and chemical similarity to polypeptides normally degraded by papain.
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| - | One example of a papain inhibitor is cystatin. According to this model, the N terminal of the cystatin interacts with the active site and the S1-S3 sites of papain. At the same time, two hairpin loops bind to the S1’-S2’ sites. The interaction between systatin and papain can be seen below. The inactivation of the cysteine proteases, including papain, occurs by competitive, noncovalent, reversible inhibition.
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| - | Another example of a papain inhibitor is <scene name='Sandbox_39/Inhibitor_revised/1'>ZLFG-DAM</scene>, a diazomethylketone inhibitor. As shown in the Jmol to the right, the methylene carbon atom of the inhibitor (shown as a grey sphere), is covalently bound to the Cys-25 of papain. The hydrophobic S2 pocket is occupied by the inhibitor's P2 side chain, shown as a pink chain. Extensive hydrogen bonding and hydrophobic interactions are responsible for the interaction of the inhibitor with the enzyme.
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| - | Several other molecules have been shown to have protease inhibitory actions. Leupeptin is a naturally-occuring, microbial protease inhibitor. Shown below, sitting in the active site of papain, this molecule contains an arginine residue at its C-terminus that is essential for its inhibitory action.
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| - | Antipain, a protease specific to papain and trypsin, is a microbial product isolated from actinomycetes. This inhibitor contains a catalytic aldehyde and acts like leupeptin. Antipain is often used in typical protease inhibitor cocktails.
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| - | [[Image:Leupeptin.jpg]]
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| - | <ref>Image from:
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| - | http://www.rcsb.org/pdb/explore/explore.do?structureId=1POP</ref>]]
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| - | ----
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| - | ----
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| - | == References ==
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| - | 1. Image from: http://upload.wikimedia.org/wikipedia/commons/5/5c/Cysteinprotease_Reaktionsmechanismus.svg
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| - | 2. Image from: http://en.wikipedia.org/wiki/Papain
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| - | 3. Hans-Hartwig Otto, and Tanja Schirmeister (1997) Cysteine Proteases and Their Inhibitors. Chemical Reviews. No. 97, 133-171.
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| - | 4. Image from:
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| - | 5. Image from: http://linus.chem.ku.edu/hewlett/Chem188/Enzyme/enzyme_background.htm
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| - | 6. Image from: http://www.rcsb.org/pdb/explore/jmol.do?structureId=9PAP&bionumber=1
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| - | 7. Image from: http://www.rcsb.org/pdb/explore/explore.do?structureId=1POP
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Adenylate kinase, commonly known as ADK, is a phosphotransferase enzyme. It has two important roles in various organisms. First, ADK plays an important role in nucleotide metabolism and synthesis. It also has a role in cellular energetics and homeostasis by phosphotransfer networks. Adenylate kinase catalyzes the reaction that forms ADP. The reaction is ATP + AMP = 2 ADP. In this catalyzed reaction, ADK molecules bind to AMP molecules and increase its binding affinity for ATP over other phosphate groups. However, adenylate kinase is also found in other molecules such as bacteria and yeast. ADK plays similar roles in bacteria and yeast, in that it involves cellular metabolism and energy. The following images highlight the structure of Adenylate kinase from Yersinia pestis, commonly known as yeast.
ADK is a protein made up of 214 amino acids. The of adenylate kinase can be seen here in purple with the ligand in the middle of the protein. The contains 12 alpha helices (light blue) and 7 beta sheets (yellow). The hold the protein together and give it shape. The are located towards the center of the protein, away from water. These residues are responsible for the overall conformation of the protein because of their "water fearing" nature. The are located mainly on the surface or outside of the protein where they will have contact with water and other molecules. The hydrophilic residues that are inside the enzyme are necessary for the enzyme to open its active site and allow substrate in. The is located in the center of protein. The ligand has six that allow it to bind to ADK's target protein (catalytic residues shown in white within the green ligand). These catalytic residues are known as the active site. The catalytic residues consist of arginine, aspartic acid, and lysine. The because it is located in the center of the protein (ligand is orange and water is displayed by space filling model). It is important to be shielded from water so that nothing interferes with the active site of the protein. The waters that are found in the interior are a result of the hydrophobic residues that are also found there. The ligands and catalytic residues of adenylate kinase are highly conserved throughout various organisms. This conservation indicates their importance to the protein's enzymatic function.
