Papain
From Proteopedia
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These two subunits are held together with <scene name='Papain/Armcrossing/1'>"arm" linkage</scene>, where each protein domain holds the opposite domain. In papain's case the "arm" crossing primarily occurs on or near the surface.<ref>[http://kinemage.biochem.duke.edu/teaching/anatax/html/anatax.2i.html] Jane S. Richardson</ref> It is between these two domains that the <scene name='Papain/9pap_bindingpocket_wrtdomains/4'>substrate binding pocket</scene> is situated.<ref>[http://books.google.com/books?hl=en&lr=&id=fk1hbZdPTEgC&oi=fnd&pg=PA79&dq=aromatic+residues+in+papain&ots=L8SvlkQaZU&sig=xZ2l8kj52PD7DzuiAQ1zah0CU2M#v=onepage&q=aromatic%20residues%20in%20papain&f=false] The Structure of Papain </ref> The two domains interact with one another via hydrophobic interactions, <scene name='Papain/Twodomainshbonds/2'>hydrogen bonds</scene> (shown in white), and electrostatic interactions in this cleft. For example, <scene name='Papain/Sk_domain_contact_valine/2'>Valine-32</scene> from the L Domain hydrophobically interacts with the carbon atoms on residues Lys174, Ala162 and Pro129 of the R Domain. <scene name='Papain/Sk_domain_contact_glutamine/3'>Gln19</scene> hydrogen bonds multiple times with the oxygen atoms of Ser176 and also with the oxygen atom on Tyr88. Electrostatic interactions are seen between <scene name='Papain/Sk_domain_contact_glu_and_lys/2'>Glu35 and Lys174</scene> where the carboxyl group of Glu35 forms an ionic bond with the ammonia group of the Lys174 residue. The <scene name='Papain/Twodomainsallncbonds/1'>sum total</scene> of interactions within the cleft between the two domains ensures that the lobes do not move with respect to one another. <ref> http://books.google.com/books?hl=en&lr=&id=fk1hbZdPTEgC&oi=fnd&pg=PA79&dq=aromatic+residues+in+papain&ots=L8SvlkQaZU&sig=xZ2l8kj52PD7DzuiAQ1zah0CU2M#v=onepage&q=aromatic%20residues%20in%20papain&f=false </ref> | These two subunits are held together with <scene name='Papain/Armcrossing/1'>"arm" linkage</scene>, where each protein domain holds the opposite domain. In papain's case the "arm" crossing primarily occurs on or near the surface.<ref>[http://kinemage.biochem.duke.edu/teaching/anatax/html/anatax.2i.html] Jane S. Richardson</ref> It is between these two domains that the <scene name='Papain/9pap_bindingpocket_wrtdomains/4'>substrate binding pocket</scene> is situated.<ref>[http://books.google.com/books?hl=en&lr=&id=fk1hbZdPTEgC&oi=fnd&pg=PA79&dq=aromatic+residues+in+papain&ots=L8SvlkQaZU&sig=xZ2l8kj52PD7DzuiAQ1zah0CU2M#v=onepage&q=aromatic%20residues%20in%20papain&f=false] The Structure of Papain </ref> The two domains interact with one another via hydrophobic interactions, <scene name='Papain/Twodomainshbonds/2'>hydrogen bonds</scene> (shown in white), and electrostatic interactions in this cleft. For example, <scene name='Papain/Sk_domain_contact_valine/2'>Valine-32</scene> from the L Domain hydrophobically interacts with the carbon atoms on residues Lys174, Ala162 and Pro129 of the R Domain. <scene name='Papain/Sk_domain_contact_glutamine/3'>Gln19</scene> hydrogen bonds multiple times with the oxygen atoms of Ser176 and also with the oxygen atom on Tyr88. Electrostatic interactions are seen between <scene name='Papain/Sk_domain_contact_glu_and_lys/2'>Glu35 and Lys174</scene> where the carboxyl group of Glu35 forms an ionic bond with the ammonia group of the Lys174 residue. The <scene name='Papain/Twodomainsallncbonds/1'>sum total</scene> of interactions within the cleft between the two domains ensures that the lobes do not move with respect to one another. <ref> http://books.google.com/books?hl=en&lr=&id=fk1hbZdPTEgC&oi=fnd&pg=PA79&dq=aromatic+residues+in+papain&ots=L8SvlkQaZU&sig=xZ2l8kj52PD7DzuiAQ1zah0CU2M#v=onepage&q=aromatic%20residues%20in%20papain&f=false </ref> | ||
- | In addition to hydrophobic residues, papain contains a variety of <scene name='Papain/Sk_polar_residues/2'>polar residues</scene>, some carrying a <scene name='Papain/Sk_acidic_residues/2'>negative charge</scene>, shown in gray at physiological pH, and are therefore acidic; others a <scene name='Papain/Sk_basic_residues/2'>positive charge</scene>, shown in purple, and are therefore basic. The rest of the <scene name='Papain/Sk_basic_residues/3'>polar residues</scene>, shown in a light gray, are neutral. As expected, the charged <scene name='Papain/Termini/ | + | In addition to hydrophobic residues, papain contains a variety of <scene name='Papain/Sk_polar_residues/2'>polar residues</scene>, some carrying a <scene name='Papain/Sk_acidic_residues/2'>negative charge</scene>, shown in gray at physiological pH, and are therefore acidic; others a <scene name='Papain/Sk_basic_residues/2'>positive charge</scene>, shown in purple, and are therefore basic. The rest of the <scene name='Papain/Sk_basic_residues/3'>polar residues</scene>, shown in a light gray, are neutral. As expected, the charged <scene name='Papain/Termini/2'>termini</scene> face outward due to their hydrophilic nature. |
Papain's secondary structure is composed of 21% <scene name='Papain/Ke_betasheets/1'>beta sheets</scene> (45 residues comprising 17 sheets) and 25% <scene name='Papain/Ke_alphahelices/1'>alpha helices</scene> (51 residues comprising 7 helices). The rest of the residues, accounting for over 50% of the enzymes structure, make up ordered non-repetative sequences.<ref name="RSCB PDB">http://www.rcsb.org/pdb/explore/explore.do?structureId=9PAP</ref> These secondary structures may be traced from the N- to C-terminus by means of <scene name='Papain/Lm_elemental/1'>differential coloration</scene>. As shown in this scene, the red end begins the protein at the N-terminus, and can be traced through the colors of the rainbow to the blue end at the C-terminus. These secondary structures form as a result of favorable hydrogen bonding interactions within the polypeptide backbone. Meanwhile, secondary structures are kept in place by hydrophobic interactions and hydrogen bonds between sidechains of adjacent structures. For example, the <scene name='Papain/Papain_sam_centralhelix/1'>first helix</scene> (residues 25-42) is maintained as a result of <scene name='Papain/Papain_sb_helix1_hbonds/1'>hydrogen bonds</scene> between backbone carbonyl atoms and the hydrogen on the amide nitrogen four residues away. However, <scene name='Papain/Papain_sam_nohbondswhelix1/1'>no hydrogen bonds</scene> are present between this helix and the rest of the protein, suggesting that this helix is coordinated primarily by hydrophobic interactions. This is reasonable given its central location in the enzyme. As expected, the helix contains many <scene name='Papain/Papain_sb_helix1_hydrophobic/1'>hydrophobic residues</scene> (red residues are hydrophilic). | Papain's secondary structure is composed of 21% <scene name='Papain/Ke_betasheets/1'>beta sheets</scene> (45 residues comprising 17 sheets) and 25% <scene name='Papain/Ke_alphahelices/1'>alpha helices</scene> (51 residues comprising 7 helices). The rest of the residues, accounting for over 50% of the enzymes structure, make up ordered non-repetative sequences.<ref name="RSCB PDB">http://www.rcsb.org/pdb/explore/explore.do?structureId=9PAP</ref> These secondary structures may be traced from the N- to C-terminus by means of <scene name='Papain/Lm_elemental/1'>differential coloration</scene>. As shown in this scene, the red end begins the protein at the N-terminus, and can be traced through the colors of the rainbow to the blue end at the C-terminus. These secondary structures form as a result of favorable hydrogen bonding interactions within the polypeptide backbone. Meanwhile, secondary structures are kept in place by hydrophobic interactions and hydrogen bonds between sidechains of adjacent structures. For example, the <scene name='Papain/Papain_sam_centralhelix/1'>first helix</scene> (residues 25-42) is maintained as a result of <scene name='Papain/Papain_sb_helix1_hbonds/1'>hydrogen bonds</scene> between backbone carbonyl atoms and the hydrogen on the amide nitrogen four residues away. However, <scene name='Papain/Papain_sam_nohbondswhelix1/1'>no hydrogen bonds</scene> are present between this helix and the rest of the protein, suggesting that this helix is coordinated primarily by hydrophobic interactions. This is reasonable given its central location in the enzyme. As expected, the helix contains many <scene name='Papain/Papain_sb_helix1_hydrophobic/1'>hydrophobic residues</scene> (red residues are hydrophilic). |
Revision as of 01:14, 5 April 2012

Papain belongs to an extended family of aminopeptidases, dipeptidyl peptidases, endopeptidases, and other enzymes having both exo- and endo-peptidase activity. The inactivated zymogen with N-terminal propeptide regions - providing stability in alkaline environments and enabling proper folding - is activated through removal of the propeptide regions. [2] The protein is primarily secreted with its pro-region enabling transport from zymogen to lysosome through membrane association and mediation. [3]
Papain. Meat tenderizer. Old time home remedy for insect, jellyfish, and stingray stings[4]. Who would have thought that a sulfhydryl protease from the latex of the papaya fruit, Carica papaya and Vasconcellea cundinamarcensis would have such a practical application beyond proteopedia?
Papain made its first appearance in the Calcutta Medical Journal entitled “The Solvent Action of Papaya Juice on Nitrogenous Articles of Food” when G.C Roy was investigating the enzyme in 1873. In the late 19th century, Wurtz and Bouchut dubbed the partially purified enzyme "papain." [5] At the time, it was viewed as a proteolytically active constituent in the latex of tropical papaya fruit. [6] As separation and purification techniques improved, pure papain was able to be isolated. In becoming the second enzyme to attain an X-ray crystallized structure and the first cysteine protease to behold an identifiable structure, papain fueled greater advances in enzymatic studies. [7]
Papain is a 23.4 kDa, 212 residue cysteine protease, also known as papaya proteinase I, from the peptidase C1 family (E.C. 3.4.22.2).[8][9] It is the natural product of the Papaya(Carica papaya)[10], and may be extracted from the plant's latex, leaves and roots. [11]. Papain displays a broad range of functions, acting as an endopeptidase, amidase, and esterase,[11] with its optimal activity values for pH lying between 6.0 and 7.0, and its optimal temperature for activity is 65 °C. Its pI values are 8.75 and 9.55, and it is best visualized at a wavelength of 278 nm. [10]
Papain's enzymatic use was first discovered in 1873 by G.C. Roy who published his results in the Calcutta Medical Journal in the article, "The Solvent Action of Papaya Juice on Nitrogenous Articles of Food." In 1879, papain was named officially by Wurtz and Bouchut, who managed to partially purify the product from the sap of papaya. It wasn't until the mid-twentieth century that the complete purification and isolation of papain was achieved. In 1968, Drenth et al. determined the structure of papain by x-ray crystallography, making it the second enzyme whose structure was successfully determined by x-ray crystallography. Additionally, papain was the first cysteine protease to have its structure identified.[11] In 1984, Kamphuis et al. determined the geometry of the active site, and the three-dimensional structure was visualized to a 1.65 Angstrom solution.[12] Today, studies continue on the stability of papain, involving changes in environmental conditions as well as testing of inhibitors such as phenylmethanesulfonylfluoride (PMSF), TLCK, TPCK, aplh2-macroglobulin, heavy metals, AEBSF, antipain, cystatin, E-64, leupeptin, sulfhydryl binding agents, carbonyl reagents, and alkylating agents.[11]
Contents |
Structure
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Inhibitors
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Common Uses
Medicinal
Papain has been used for a plethora of medicinal purposes including treating inflammation, shingles, diarrhea, psoriasis, parasites, and many others.[26] One major use is the treatment of cutaneous ulcers including diabetic ulcers and pressure ulcers.[27] Pressures ulcers plague many bed bound individuals and are a major source of pain and discomfort. Two papain based topical drugs are Accuzyme and Panafil, which can be used to treat wounds like cutaneous ulcers.[28]
A recent New York Times article featured papain and other digestive enzymes.[29] With the number of individuals suffering from irritable bowel syndrome and other gastrointestinal issues, many people are turning toward natural digestive aid supplements like papain. The author even talks about the use of papain along with a pineapple enzyme, bromelain, in cosmetic facial masks. Dr. Adam R. Kolker (a plastic surgeon) is quoted in the article saying that "For skin that is sensitive, enzymes are wonderful." He bases these claims off the idea that proteases like papain help to break peptide bonds holding dead skin cells to the live skin cells.[30]
Commercial and Biomedical
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. This non-specific nature of papain's hydrolase activity has led to its use in many and varied commercial products. It is often used as a meat tenderizer because it can hydrolyze the peptide bonds of collagen, elastin, and actomyosin. It is also used in contact lens solution to remove protein deposits on the lenses and marketed as a digestive supplement. [31] Finally, papain has several common uses in general biomedical research, including a gentle cell isolation agent, production of glycopeptides from purified proteoglycans, and solubilization of integral membrane proteins. It is also notable for its ability to specifically cleave IgG and IgM 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.[11]
Fun Trivia
Remember the 2002 SARS (Severe Acute Respiratory Syndrome) epidemic that placed global health, particularly in Southeast Asia, in a precarious state? On-going research is happening to further understand the mechanisms of this coronavirus, so that future steps can be taken for prevention. Its been found that the replication of RNA for this virus is mediated by two viral proteases that have many papain-like characteristics. [32]
Despite a low percentage of sequence identities, inhibition and sequence analyses have increasingly been drawing parallels between L proteinases, that involve the foot-and-mouth disease virus and equine rhinovirus 1, and papain. With a similar overall fold to papain and identifiable regions that resemble the five alpha-helices and seven beta-sheets of papain, L proteinases of foot-and-mouth disease virus and of equine rhinovirus 1 reveal a mode of operation that is very papain like. [33]
References
- ↑ [1] Papaya's Nutrition Facts
- ↑ Rawlings ND, Barrett AJ. Families of cysteine peptidases. Methods Enzymol. 1994;244:461-86. PMID:7845226
- ↑ Yamamoto Y, Kurata M, Watabe S, Murakami R, Takahashi SY. Novel cysteine proteinase inhibitors homologous to the proregions of cysteine proteinases. Curr Protein Pept Sci. 2002 Apr;3(2):231-8. PMID:12188906
- ↑ [2] Ameridan International
- ↑ Menard and Storer 1998
- ↑ Wurtz and Bouchut 1879
- ↑ Drenth J, Jansonius JN, Koekoek R, Swen HM, Wolthers BG. Structure of papain. Nature. 1968 Jun 8;218(5145):929-32. PMID:5681232
- ↑ [3] Uniprot
- ↑ 9.0 9.1 [4] 9PAP PDB
- ↑ 10.0 10.1 [5] Sigma Aldrich
- ↑ 11.0 11.1 11.2 11.3 11.4 [6] Worthington
- ↑ 12.0 12.1 Kamphuis IG, Kalk KH, Swarte MB, Drenth J. Structure of papain refined at 1.65 A resolution. J Mol Biol. 1984 Oct 25;179(2):233-56. PMID:6502713
- ↑ [7] Jane S. Richardson
- ↑ [8] The Structure of Papain
- ↑ http://books.google.com/books?hl=en&lr=&id=fk1hbZdPTEgC&oi=fnd&pg=PA79&dq=aromatic+residues+in+papain&ots=L8SvlkQaZU&sig=xZ2l8kj52PD7DzuiAQ1zah0CU2M#v=onepage&q=aromatic%20residues%20in%20papain&f=false
- ↑ http://www.rcsb.org/pdb/explore/explore.do?structureId=9PAP
- ↑ http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html
- ↑ [9] University of Maine
- ↑ [10]Harrison, M.J., N.A. Burton, and I.H. Hillier. 1997. Catalytic Mechanism of the Enzyme Papain: Predictions with a Hybrid Quantum Mechanical/Molecular Mechanical Potential. J. Am. Chem. Soc. 119: 12285-12291
- ↑ [11] Schröder, E., C. Phillips, E. Garman, K. Harlos, C. Crawford. 1997. X-ray crystallographic structure of a papain-leupeptin complex. FEBS Letters 315: 38-42
- ↑ LaLonde JM, Zhao B, Smith WW, Janson CA, DesJarlais RL, Tomaszek TA, Carr TJ, Thompson SK, Oh HJ, Yamashita DS, Veber DF, Abdel-Meguid SS. Use of papain as a model for the structure-based design of cathepsin K inhibitors: crystal structures of two papain-inhibitor complexes demonstrate binding to S'-subsites. J Med Chem. 1998 Nov 5;41(23):4567-76. PMID:9804696 doi:10.1021/jm980249f
- ↑ http://pubs.acs.org/doi/abs/10.1021/ci800085c
- ↑ http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2923042/?tool=pubmed
- ↑ http://www.biomedcentral.com/1472-6807/10/30
- ↑ http://www.ncbi.nlm.nih.gov/pmc/articles/PMC551902/pdf/emboj00233-0254.pdf
- ↑ http://www.webmd.com/vitamins-supplements/ingredientmono-69-PAPAIN.aspx?activeIngredientId=69&activeIngredientName=PAPAIN
- ↑ http://www.pbm.va.gov/Clinical%20Guidance/Drug%20Monographs/Papain%20Urea.pdf
- ↑ http://www.pbm.va.gov/Clinical%20Guidance/Drug%20Monographs/Papain%20Urea.pdf
- ↑ http://www.nytimes.com/2012/02/23/fashion/enzymes-once-sidelined-try-to-grab-the-spotlight.html
- ↑ http://www.nytimes.com/2012/02/23/fashion/enzymes-once-sidelined-try-to-grab-the-spotlight.html
- ↑ http://www.webmd.com/vitamins-supplements/ingredientmono-69-PAPAIN.aspx?activeIngredientId=69&activeIngredientName=PAPAIN
- ↑ Barretto N, Jukneliene D, Ratia K, Chen Z, Mesecar AD, Baker SC. The papain-like protease of severe acute respiratory syndrome coronavirus has deubiquitinating activity. J Virol. 2005 Dec;79(24):15189-98. PMID:16306590 doi:10.1128/JVI.79.24.15189-15198.2005
- ↑ Skern T, Fita I, Guarne A. A structural model of picornavirus leader proteinases based on papain and bleomycin hydrolase. J Gen Virol. 1998 Feb;79 ( Pt 2):301-7. PMID:9472614
Reference
- Wang J, Xiang YF, Lim C. The double catalytic triad, Cys25-His159-Asp158 and Cys25-His159-Asn175, in papain catalysis: role of Asp158 and Asn175. Protein Eng. 1994 Jan;7(1):75-82. PMID:8140097
- Kamphuis IG, Kalk KH, Swarte MB, Drenth J. Structure of papain refined at 1.65 A resolution. J Mol Biol. 1984 Oct 25;179(2):233-56. PMID:6502713
3D Structures of Papain
3LFY, 1KHP, 1KHQ, 1CVZ, 1BQI, 1BP4, 1PPN, 1PPP, 1PIP, 1POP, 1PE6, 9PAP, 1PPD, 1PAD, 2PAD, 4PAD, 5PAD, 6PAD- Carica papaya
3IMA - Colocasia esculenta
1STF - Homo sapiens
2CIO - Trypanosoma brucei
3E1Z - Trypanosoma cruzi
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