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== '''Papain''' (PDB ID #: 9pap)==

<Structure load='Insert PDB code or filename here' size='500' frame='true' align='right' caption='Insert caption here' scene='Sandbox_31/Primary_scene/2' />

Papain is a cysteine protease (EC 3.4.22.2) that is found naturally occurring in [http://en.wikipedia.org/wiki/Carica_papaya Papaya](''Carica papaya'').<ref name="Sigma Aldrich">http://http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html</ref> Papain is extracted from the latex, leaves and roots of the papaya tree.<ref name="worthington">http://www.worthington-biochem.com/pap/default.html</ref> Thiol proteases like papain are commonly found in different fruits like pineapples, figs, and kiwis. Proteases are a group of enzymes that cleave peptide bonds of protein molecules. Papain is a protease commonly used for cell isolation, protein structural studies, cleaving antibodies, and various other research techniques.<ref name="worthington">http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html</ref> In addition to use in research, it is commonly used in industry to tenderize meats, remove skin from hides, and clean soft contact lenses.

Molecular Weight: 23,406 daltons<ref name="Sigma Aldrich">http://http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html</ref>

PI: 8.75<ref name="worthington">http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html</ref>

Optimal pH: 6.0-7.0<ref name="Sigma Aldrich">http://http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html</ref>

==Structure==

Papain is formed from a peptide that is 212 amino acids in length.<ref>http://www.pdb.org/pdb/explore/explore.do?structureId=9PAP</ref> It consists of 25% <scene name='Sandbox_31/Alpha_helices/4'>Alpha helices</scene> and 21% <scene name='Sandbox_31/Beta_sheet/2'>Beta Sheet</scene>. The structure also consists of three <scene name='Sandbox_31/Disulfide/3'>Disulfide bonds</scene> between <scene name='Sandbox_31/Cys_with_labels/2'>Cysteine Residues</scene>. The disulfide bonds exist between Cys 63 and Cys 22, Cys 200 and Cys 153, and Cys 56 and Cys 95. Another important residue to the structure is Cysteine 25 which has a sulfhydryl group that plays a major role in the active site activity[[http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html]].

The <scene name='Sandbox_31/Hydrophobic_residues/2'>Hydrophobic Residues</scene> in papain are primarily located toward the inside of the enzyme or paired by hydrophobic interactions with other hydrophobic residues to exclude water. The <scene name='Sandbox_31/Polar_residues/1'>Polar Residues</scene> are located facing the exterior, in the active site, or paired with other charged residues. The polar residues seen in orange predominantly fill the space surrounding the exterior of the enzyme. This is because they are hydrophillic and interact with polar water molecules. The <scene name='Sandbox_31/Positive/1'>Negatively</scene> charged residues (shown in green) and the <scene name='Sandbox_31/Basic_positive/1'>Positively</scene> charged residues (shown in red) make up the polar residues. The charged and polar residues are hydrophilic therefore do not orient themselves to exclude water or solvent. The charged <scene name='Sandbox_31/Termini/4'>Termini</scene> face outward due to their hydrophilic nature.

 

 

<scene name='Sandbox_31/Leupeptin/1'>Leupeptin</scene> is a commonly studied inhibitor of proteases (seen in the Jmol as ball and stick model). It inhibits by binding and interacting with the active site which allows it to block the enzyme's desired protein substrate. There are many <scene name='Sandbox_31/1popligand_contacts/1'>Residues</scene> that interact with Leupeptin in the active site. The predominant interaction is from hydrophobic interactions between Leupeptin and <scene name='Sandbox_31/1pophydrointeract/1'>active site residues</scene>. In addition to hydrophobic interactions, there are also some hydrogen bonding interactions to hold Leupeptin in the active site of papain. Leupeptin works well at blocking papain from its enzymatic duties. A recent study has shown that Leupeptin actually forms a covalent bond between its <scene name='Sandbox_31/Leupeptin_active/1'>Carbonyl Carbon</scene> and CYS 25. In addition, the residues Gln 19 and CYS 25 form <scene name='Sandbox_31/Leupeptin_active/2'>hydrogen bonds</scene>with the Leupeptin molecule

 

 

Cathepsin L is another inhibitor of the papain enzyme. Cathepsin L interacts with the <scene name='Sandbox_31/Cathepl/1'>papain residues</scene> Gln19, Cys25, Gly66, Asp158, and Trp177 by hydrogen bonding them (Cathepsin L is lime green in the model, and the papain residues active in hydrogen bonding with Cathepsin L are highlighted with yellow halos). In addition to hydrogen bonding, hydrophobic interactions exist to exclude water, allowing the papain enzyme and Cathepsin L to associate even closer. Cathepsin L plays a roll in many different diseases including malaria, leishmaniasis, Chagas' disease, African trypanosomiasis, toxoplasmosis, and amoebiasis.<ref>http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2923042/?tool=pubmed</ref>Some studies show that there is a relation between cathepsins and certain cancers, alzheimer's, and arthritis.<ref>http://www.biomedcentral.com/1472-6807/10/30</ref>

 

 

Solvents play a role as papain ligands. One such solvent is <scene name='Sandbox_31/Methanol/1'>Methanol</scene>, which can be seen as the blue molecules surrounding the papain enzyme. Along with methanol are <scene name='Sandbox_31/Water/1'>water</scene> molecules, which can be seen in red. Both methanol and water likely act to stabilize the enzyme structure through hydrogen bonding interactions.

The active site of papain has a <scene name='Sandbox_31/Activesite/1'>"Catalytic Triad"</scene>including CYS 25, HIS 159, and ASN 175.

[[Image:papain2.jpg|150px|left|thumb|Active Site of Papain<ref>

http://peds.oxfordjournals.org/content/7/1/75.abstract</ref>]]

This triad interacts with the substrate to catalyze the reaction. The sulfhydryl group on CYS 25 plays the key role in the mechanism, which is why papain is considered a thiol protease. The sulfur from CYS 25 attacks the backbone amine on the substrate forming a tetrahedral intermediate. Next, the carbonyl is reformed and the carbon nitrogen bond is broken. A water associated with a nitrogen on HIS 159 then attacks the carbonyl forming a second tetrahedral intermediate. The carbonyl then reforms breaking the carbon-sulfur bond. This leaves a carboxy group on the end of one piece of the substrate and an amino group on the end of the other piece.<ref>http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html

</ref> The end product is two protein fragments.

'''Oxyanion Hole'''

 

The <scene name='Sandbox_31/Oxhole/1'>Oxyanion hole</scene> plays an important role in binding and holding the substrate protease. The papain oxyanion hole consists of CYS 25 and Gln 19. The carbonyl of the peptide bond to be cleaved is bound in the oxyanion hole with CYS 25. The oxyanion hole stabilized the enzyme substrate complex.

 

[[Image:mech.jpg|400px|right|thumb|Proteolytic Mechanism of the Papain Enzyme with substrate<ref>http://chemistry.umeche.maine.edu/CHY431/Peptidase10.html</ref>]]

 

 

'''For a Video Explanation of the Cysteine protease mechanism''' [http://www.youtube.com/watch?v=-6WSmDjbuzA Click Here].

 

 

 

 

 

 

 

 

 

[[Image:papayas.jpg|150px|left|thumb|Papaya<ref>http://dailyfitnessmagz.com/2011/03/papayas-nutrition-facts/</ref>]]

 

 

http://www.pdb.org/pdb/explore/remediatedSequence.do?structureId=9PAP

 

http://faculty.csusm.edu/lcohen/index.html