Laccase in general is an enzyme which contains multiple coppers inside. (Mainly, it is in a state of Cu2+ or Cu+). Some alternative names are p-diphenol oxidase, urishiol oxidase, or urushiol oxidase. (1) Each subunit contains type one, type two and type three copper sites or each active protein site needs a minimum of four copper atoms. Usually, a mononuclear type 2 copper atom and a binuclear type 3 pair together are called trinuclear cluster. (2) That is, the two types of copper atoms stay close to each other and have similar functions within the whole enzyme. Thus, laccase belongs to class of blue multi-copper oxidase. (3) In addition, laccase is the oxidase which has low specificity in its one-electron oxidation with various substrates. Inside the subunit, the type 1 copper atom is capable of accepting electrons from a reducing substrate and transfers them to the type 2-type 3 trinuclear cluster. At the time, dioxygen (or O2) are attracted to the trinuclear center and be converted to water in this region. It is the reduction of dioxygen to water. (3) Laccase has a capacity of oxidizing a large number of substrates. For example, it can act on some phenol compounds. Thus, laccase is often said to catalyze the degradation of lignin since lignin contains many phenolic groups. (4) In this way, the function of laccase in some wood and litter degrading fungi is that they help soil organic compound turnover as well as to attribute in global carbon cycle since lignin is a very abundant polymer on Earth having around 30% of non-fossil organic carbon. It is also important in some physiological process such as pathogenesis and morphogenesis. One instance involves the pigmentation. Other than its discovery in some fungi, laccase also could be found in many plants and microorganisms. (5)
- Type 1 copper atom in the subunit of laccase is the site that accepts the electrons from reducing substrates. (6)Type one copper atom attaches to four other ligands: 2 His, 1 Cys, and a Met sulfur. However, the Met sulfur is relatively weak and the type 1 copper in the laccase actually possess no principal axis or rotation because the core of it is a trigonal planar or it has the coordination number of 3. (The copper is connected to two His and one Cys) However, it contains one α plane. Thus, it belongs to the Cs point group. That is, the only symmetry element for this compound is E and the α plane. Since type 1 copper atom belongs to the Cs point group, it is both IR and Raman active according to the character table.
- Type 2 copper atom in the laccase actually works with a binuclear type 3 pairs and so called the trinuclear cluster. The type two copper is bound to two His and an OH group. Its molecular geometry is trigonal pyramid. The only symmetry element for it is E and thus it belongs to C1 point group. However, laccase structure with absence of type 2 copper atom has been reported. No IR or Raman bands will be observed for type 2 copper.
- Two type 3 copper atoms are connected together by an OH and each of the type 3 copper atom is connected to other three His groups. The binuclear type 3 copper atom pair with type 2 copper is the trinuclear cluster in the laccase and can reduce dioxygen to 2 water molecules. (6) Totally they (type 2 and 3) have eight His ligands connected to them, but if only focus on one type 3 copper as the metal center, the type 3 copper is connected to three His group and one very large group which could be viewed as a R group here. Since this R group is heavier than the three His ligands, type 3 copper metal with its ligands belongs a C3v point group, where the principal axis lies at the copper-R axis. Since the focus is on one of the type 3 copper and it belongs to C3v point group, the character chows that the molecule is both IR and Raman active.
Overall, the subunit region of the laccase is not symmetric. (The four copper atoms) Thus, the only suitable point group is C1 indicating the E symmetry element. No IR or Raman will be observed. In general the laccase enzyme is asymmetric but its presence has been detected by spectrophotometer which takes advantage of some of the suitable substrates such as ABTS. In addition, the activity of the laccase could be detected by an oxygen sensor since the reaction of laccase involves the oxidation of substrates as well as the reduction of dioxygen to water. (7)
However, the actual enzymativally active form of the laccase could be composed of two subunits or three subunits. That is, it is a polymeric enzyme. (7) In the case of lignin degradation by laccase, the reaction occurs outside of the cell while some other laccase enzyme reacts with a wide range of substrates inside the cell. Moreover, a test shows that laccase which performs lignin degradation in the fungus has two laccase molecules, naming A and B now. And from the relative detection of molecule oxygen in its reduction in the trinuclear cluster, it is found that two intermediates (“peroxide” and “native”) are in each of the two laccase molecules. For example, in laccase molecule B, the two type 3 copper atoms are connected by a peroxide ion while the other two type 3 copper atoms in laccase molecule A are connected by a hydroxide ion. (3)
Some of the interesting things regarding laccase is that laccase is viewed as a monomer which contains three “cupredoxin-like beta-sandwich domains.” Laccase is found to be similar to ascorbate oxidase in structure. (4) However, one difference is that laccase has a type one copper atom that are connected to four other ligands ( two his, one Cys and a Met sulfur) actually exhibits trigonal planar molecular geometry or the corresponding coordination number is 3.As a result, type-1 copper atom structure is suspected to be responsible for the high redox potential. Moreover, it is hard to detect type 2 copper atom in laccase and it results in a so called “type-2 depletion” condition which influences the type-3 copper atom in its reactivity since the ability to reduce oxygen to water requires both the type 2 and two type 3 in the trinuclear cluster and in their active forms. (8)
- “E.C 18.104.22.168” http://www.ebi.ac.uk/intenz/query?cmd=SearchEC&ec=22.214.171.124.
- “Definition of Laccase.” http://www.chemicool.com/definition/laccase.html.
- Ferraroni, M; N.M. Myasoedova; V Schmathenko.e.t. Crystal structure of a blue laccase from Lentinus tigrinus: evidences for intermediates in the molecular oxygen reductive splitting by multicopper oxidases. Bmc Struct Biol [Online], 2007, 7, pp 60.
- Ducros, V; A.M. Brzozowski; K.S. Wilson; S.H. Brown,e.t. Crystal structure of the type-2 Cu depleted laccase from Coprinus cinereus at 2.2 A resolution. Nat Struct Biol.[Online] 1998, 5, pp 310-316.
- “Laccase Project.” http://www.haraldkellner.com/html/laccase_project.html.
- Lyashenko, A.V; N.E. Zhukhlistova; A.G. Gabdoulkhakov.e.t. Purification, crystallization and preliminary X-ray study of the fungal laccase from Cerrena maxima. Acta Crystallograph Sect F Struct Biol Cryst Commun, [Online] 2006, 62, pp 954-967.
- “Laccase.” http://en.wikipedia.org/wiki/Laccase.
- Piontek, k; M. Antorini; T. Choinowski. Crystal structure of a laccase from the fungus Trametes versicolor at 1.90-A resolution containing a full complement of coppers. J Biol Chem. [Online]. 2002, 277, pp 37663-37669.