Sandbox 34

Please do NOT make changes to this Sandbox. Sandboxes 30-60 are reserved for use by Biochemistry 410 & 412 at Messiah College taught by Dr. Hannah Tims during Fall 2012 and Spring 2013. Papain Introduction spinqualitylabelsall models Papain Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Papain is a cysteine protease, also known as papaya proteinase I, from the peptidase C1 family with E.C. 3.4.22.2. It functions as a hydrolase, endopeptidase, and thiol protease. Its optimal activity values for pH lie between 6.0 and 7.0, and 65 °C as its optimal temperature for activity. Its pI values are 8.75 and 9.55. Papain is best visualized at a wavelength of 278 nm. Naturally found in the latex of the papaya fruit, one of the most common uses of papain is as a meat tenderizer because of its ability to hydrolyze esters and amides. Another common use is as a digestive aid. Papaya is commonly referenced as a preferred fruit for those suffering from gastroesophageal reflux disease due to its ability to help the the stomach with digestion of complex proteins. History 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. 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. Today, studies continue on the stability of papain, involving changes in environmental conditions, in addition to 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. Structure Papain is a relatively simple enzyme. It consists of only one chain of 212 residues with three disulfide bonds, illustrated in yellow. A modified cysteine residue with a sulfhydryl group, cysteine sulfonic acid, is necessary for the activity of the enzyme. In 9PAP, the sulfhydryl group has been oxidized. Papain contains many hydrophobic regions illustrated in magenta. Ligands Papain binds many methanol molecules via hydrogen bonding. Specificity Papain digests a large variety of proteins, with a very broad specificity. Its active siteconsists of the residues cysteine-25, histidine-159, and asparagine-175. It cleaves the peptide bonds of basic amino acids, leucine and glycine by nucleophilic attack with its sulfhydryl group on cysteine-25 . It also hydrolyzes esters and amides. It prefers amino acids that bear large hydrophobic side chains at the P2 position, and will not accept valine at the P1' position. Catalytic Mechanism The mechanism of cysteine proteases is very similar to that of serine proteases. The sulfhydryl group on cysteine executes a nucleophilic attack on the peptide bond of the protein it wishes to cleave. Opening up the carbonyl, the sulfhydryl group of CYS-25 is stabilized by HIS-159. As the carbonyl reforms, the peptide bond is broken, leaving the amide group to fend for itself. General mechanism of papain catalysis. Inhibitors There are many inhibitors for papain because of its broad specificity. The inhibition of papain is usually due to active site restriction. An interesting inhibitor is that of Clik148. This inhibitor uses both prime and non-prime sites to inhibit papain. References External References Wikipedia