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=  Laccase  = Laccases (EC 1.10.3.2) are copper-containing oxidase enzymes found in many plants, fungi, and microorganisms. Laccases act on phenols and similar molecules, performing one-electron oxidations, which remain poorly defined. It is proposed that laccases play a role in the formation of lignin by promoting the oxidative coupling of monolignols, a family of naturally occurring phenols. Other laccases, such as those produced by the fungus Pleurotus ostreatus, play a role in the degradation of lignin, and can therefore be classed as lignin-modifying enzymes.

Laccases require oxygen as a second substrate for their enzymatic action.

Spectrophotometry can be used to detect laccases, using the substrates ABTS, syringaldazine, 2,6-dimethoxyphenol, and dimethyl-p-phenylenediamine. Activity can also be monitored with an oxygen sensor, as the oxidation of the substrate is paired with the reduction of oxygen to water.

Laccase was first studied by Gabriel Bertrand in 1894 in the sap of the Japanese lacquer tree, where it helps to form lacquer, hence the name laccase.

Laccases can catalyze ring cleavage of aromatic compounds.

Copper Binding
The copper bound by laccase is bound in several sites: type 1, type 2, and/or type 3. The ensemble of types 2 and 3 copper is called a trinuclear cluster (see figure). Type 1 copper is available to action of solvents, such as water. It can be displaced by mercury, substituted by cobalt or removed via a copper complexone. Removal of type 1 copper causes a decrease in laccase activity. Cyanide can remove all copper from the enzyme, and re-embedding with type 1 and type 2 copper has been shown to be impossible. Type 3 copper, however, can be re-embedded back into the enzyme.

Inhibition
Laccase can be inhibited by small ions such as: azide, halides, cyanide, and fluoride. These ions bind to type 2 and type 3 copper and disrupts electron transfer via copper centers, therefore reduces activity. Metal ions, fatty acids, hydroxyglycine, and kojic acid can also inhibit laccase by causing amino acid residue changes, conformational changes or copper chelation.[6]

Applications and potential utility
Laccases have been examined as the cathode in enzymatic biofuel cells. They can be paired with an electron mediator to facilitate electron transfer to a solid electrode wire.[7] Laccases are some of the few oxidoreductases commercialized as industrial catalysts. The enzymes can be used for textile dyeing/textile finishing, wine cork making, teeth whitening, and many other industrial, environmental, diagnostic, and synthetic uses.[8] Laccases can be used in bioremediation. Protein ligand docking can be used to predict the putative pollutants that can be degraded by laccase.[9]

Laccase is used widely in the following industries:


 * Textile Industry
 * Food Industry
 * Pharmaceutical Industry
 * Other[10]

Activity in Wheat Dough
Laccases have the potential to cross link food polymers such as proteins and nonstarch polysaccharides in dough. In non starch polysaccharides, such as arabinoxylans (AX), laccase catalyzes the oxidative gelation of feruloylated arabinoxylans by dimerization of their ferulic esters.[11] These cross links have been found to greatly increased the maximum resistance and decreased extensibility of the dough. The resistance was increased due to the crosslinking of AX via ferulic acid and resulting in a strong AX and gluten network. Although laccase is known to cross link AX, under the microscope it was found that the laccase also acted on the flour proteins. Oxidation of the ferulic acid on AX to form ferulic acid radicals increased the oxidation rate of free SH groups on the gluten proteins and thus influenced the formation of S-S bonds between gluten polymers.[12] Laccase is also able to oxidize peptide bound tyrosine, but very poorly.[12] Because of the increased strength of the dough, it showed irregular bubble formation during proofing. This was a result of the gas (carbon dioxide) becoming trapped within the crust and could not diffuse out (like it would have normally) and causing abnormal pore size.[11] Resistance and extensibility was a function of dosage, but at very high dosage the dough showed contradictory results: maximum resistance was reduced drastically. The high dosage may have caused extreme changes in structure of dough, resulting in incomplete gluten formation. Another reason is that it may mimic overmixing, causing negative effects on gluten structure. Laccase treated dough had low stability over prolonged storage. The dough became softer and this is related to laccase mediation. The laccase mediated radical mechanism creates secondary reactions of FA-dervived radicals that result in breaking of covalent linkages in AX and weakening of the AX gel.[11]

Use in Food Industry
The hazing effect is a quality defect in beer. It is characterized by “cloudiness” in the final product. Laccase can be added to the wort or at the end of the process to remove the polyphenols that may still remain in beer. The polyphenol complexes, formed by laccases, can be separated via filtration and removes probability of the hazing effect from occurring.

Laccase can also remove excess oxygen in beer and increase the storage life of beer.

In fruit juices such as apple and grape, excess oxidation of phenolics causes negative effects on the taste, color, odour and mouthfeel. Laccase has been proposed to delay the oxidation of polyphenols and stabilize the juice.

Origin and Discovery
Laccase enzymes were first discovered in the lacquer of a Japanese tree called Toxicodendron veniciflumm (Rodriguez-Couto). Laccase can be found in fungi, plants, bacteria and insects.

Background
Laccase enzymes was first discovered in the lacquer of a Japanese tree called Toxicodendron veniciflumm. Laccase can be found in fungi, plants, bacteria and insects. Laccases participate in the break down of polymers and cleaving aromatic rings. This is found in plants (cabbage,turnips and apples) to name a few and fungi such as Ascomycetes and Deuteromycetes. Lately there has been some strains that are found in bacteria S.cyaneus. Laccases have a role in the food and industrial industry because they are used to make paper and textile. Laccases are used to remove pollution from the earth and found to clean up ionic spills and used to remove peptides. Laccases are also found in cosmetics such as eyeshadows and highlighter and other beauty products. Laccases can be used as a catalyst from drugs that can cure cancer.

Laccase has been found to inhibit HIV-1 reverse transcriptase. Several studies have been done to isolate and look further into the properties and applications of different strands of Laccases, in which some of those strands have been found to inhibit HIV-1 RT.

Mechanisms
Laccase catalysis occurs because of the decrease in the ratio of oxygen to water. Laccase contains 4 copper atoms. T1: gives a blue hue when the enzyme absorbed around 600nm, Type 2 does not give a detectably color and T3: has binuclear configuration that gives a weak absorbance.

Laccase a polyphenol oxidase. The oxygen is reduced to water and is accompanied by oxidation of one electron with from a wide of aromatic compounds. '''(Bourbonnais)  During the reaction of laccase, diphenol (hydroquinone undergoes a one-electron oxidation to form an oxygen centered free radical. The species can then be converted to the quinone in a second enzyme-catalyzed step or by non-spontaneous disproportion. (Thurston, Christopher)'''

Current Studies
Recent study conducted on Laccase pretreatment for agrofood wastes valorization. The experiment conducted on agrofood wastes ( apple pomace, potato peels and coffee grounds) to test its effectiveness in hydrolysis when lignin is removed. Agrofood wastes are important for their production of biofuels and chemicals but their conversion is limited by the presence of lignin, a organic polymer found in cell wall of plants. The experiment used laccase enzyme from Pleurotus ostreatus to help remove the lignin polymer in plans in order to measure the effectiveness of the hydrolysis of agro foods to produce beneficial biofuels. Results revealed 83% hydrolysis yields for apple pomace when using laccase As a result a sequential protocol was developed to effectively produce biofuels. ('''Rodriguez-Couto, Susan) ''' There was another study done by Zhao et al. that aimed to derive Laccase from a mushroom (Coprinus comatus), or CCL as described in study, and look further into its properties as well as its applications. The researchers found that the native Coprimus comatus Laccase is a monomeric protein. CCL was also shown to stop proliferative activity in certain tumor cell lines (HepG2 and MCF7) and it was also shown to inhibit HIV1-RT, which shows that CCL is a pathogenic protein. The pH and temperature at which CCL was at its most optimum was also shown to be at 2.0 and 60oC respectfully.

Properties
Laccase has three forms monomeric,monomeric, dimeric and tetrameric protein. Glycosylation allows for copper rendition and temperature and secretion if laccase. The optimal temperature is about 25 degrees celsius.

Forms for different strains of Laccase
C. comatus - monomeric protein

Phellinus ribis - dimeric protein

Gaeumannomyces graminis - quadruple protein

Optimal pH for different strains of Laccase
C. comatus - 2.0

A. cylindracea - 5.2

P. nebrodensis - 5.0

L. ventriosospora - 4.0

A. blazei - 2.3

R. virescens - 2.2