From Proteopedia

Revision as of 02:41, 14 November 2011

Papain

Introduction

is a cysteine protease that is stable and active under a wide range of conditions. The enzyme present in the leaves, latex, roots, and fruit of the papaya plant (Carica papaya).^[1] The papain proteins are synthesized as inactive precursors that become active within two minutes of the plant being wounded and the latex is expelled.^[2] The latex is dried and then purified to extract the active papain enzme. The enzyme was first studied and isolated in the 1960's. It has a 23.4kDa theoretical molecular weight and works at an optimum pH of 6-7 and optimum temperature of 60-70 degrees Celsius.

Structure

Papain's three-dimensional structure is at 1.65 Angstrom resolution. It consists of one polypepetide chain that is made up of 212 amino acids residues. There are three present in the enzyme that maintains the protein's structure. Papain consists of 25% alpha helices and 21% beta sheets within its . Alpha helices are shown in red and beta sheets are shown in green. The enzyme's structure has a mixture of shown in yellow and hydrophilic residues shown in purple. This corresponds to the protein's shown in orange and nonpolar residues shown in green. Papain is naturally found with many . The majority are methanol molecules because the crystallization medium is 62% (w/w) methanol in water and only 29 out of 224 solvent molecules can be regarded with any certainty as methanol molecules.^[3] Here, the enzyme's sulfhydryl group can also be seen which is necessary for the activity of the enzyme. and between the ligands and protein maintain the structural domains of papain. Water molecules are depicted as pink spheres.Twenty-one water molecules are located in contact areas between adjacent papain molecules.

Mechanism

Papain cleaves peptide bonds of basic amino acids, leucine, or glycine. It also hydrolyzes esters and amides.^[4] Its mechanism of breaking peptide bonds takes place in its . The active site is in a cleft formed by the two distinct structural domains of the protein. The active site contains a catalytic dyad of residues cysteine-25 on an alpha helix and histidine-159 on a beta sheet. The catalytic dyad is similar to the catalytic triad seen in most serine proteases. This ion pair of cysteine and histidine is the powerhouse of papain's catalytic mechanism. The other residue of the active site is asparagine-175, which helps with orientation during catalysis. For catalysis, His-159 deprotonates cys-25, then cys-25 attacks the carbonyl of the peptide as a nucleophile. A water molecule then deacylates the enzyme and releases the peptide.

Inhibition

One inhibitor of papain is leupeptin. The carbonyl carbon of this (seen in blue) is covalently bound by the Cys-25 sulphur atom of papain and is tetrahedrally coordinated. The carbonyl oxygen atom of the inhibitor faces the oxyanion hole and makes hydrogen bond contacts with Gln-19 and Cys-25.^[5] The and between papain and leupeptin keep the active site closed and the residues (shown in brown) can no longer catalyze.

Function

Papain is used as a powdered meat tenderizer because of its ability to break down tough meat fibers. It can also be used as a treatment for jellyfish and bee stings because it can break down the protein toxins in the venom.^[6] Papain can also digest immunoglobulins (antibodies) by breaking the two Fab fragments from the Fc fragment. This prevents the immunoglobulin's ability to promote agglutination.

References

1. ↑ http://www.britannica.com/EBchecked/topic/441803/papain
2. ↑ http://www.worthington-biochem.com/pap/default.html
3. ↑ http://www.pdb.org/pdb/explore.do?structureId=9PAP
4. ↑ http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/analytical-enzymes/papain.html
5. ↑ http://www.pdb.org/pdb/explore/remediatedSequence.do?structureId=1POP&params.chainEntityStrategyStr=all
6. ↑ http://en.wikipedia.org/wiki/Papain