title: Activity and stability of laccase in conjugation with chitosan.
abstract: Laccase is one of a few enzymes that can directly reduce oxygen into water under ambient conditions, while oxidizing a variety of aromatic compounds. Its conjugation with chitosan generates a pH-sensitive functional biomaterial that changes its solubility in response to pH variation. The molecular conjugation between laccase and chitosan of different molecular mass was investigated with a carbodiimide reaction to understand the mechanism of the enzyme's activity loss during conjugation. With 81-93% laccase being conjugated, a moderate activity loss (16-28% less than the initial activity) was observed in conjugation solution. A second severe activity loss (63-78% less than the conjugated activity) occurred during a cycle of phase change consisting of precipitation, centrifugation and re-dissolution of the enzyme-chitosan conjugates. The chitosan molecular size has little effect on the first moderate activity loss in the conjugation reaction, but visible effect on the substantial activity loss associated with phase change. Small chitosan molecules gave high residual activity. The conjugated laccase exhibited a high stability in the following repeated phase changes and had the same temperature and pH profile as those of free laccase. Compared to free laccase, the conjugated laccase had a similar affinity (Km), but reduced turnover (kcat) that was adversely affected with increase of molecular mass of chitosan.
authors.affiliation: Department of Molecular Bioscience and Bioengineering, Honolulu, HI 96822, USA.
authors.forename: Gary
authors.initials: G
authors.lastname: Delanoy
authors.forename: Qing
authors.initials: Q
authors.lastname: Li
authors.forename: Jian
authors.initials: J
authors.lastname: Yu
mesh_terms: D001672:Biocompatible Materials / Q000737:chemistry*; D002234:Carbodiimides / Q000737:chemistry; D048271:Chitosan / Q000737:chemistry*; D002845:Chromatography; D003300:Copper / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004798:Enzymes / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D042845:Laccase / Q000737:chemistry*; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010100:Oxygen / Q000737:chemistry; D011485:Protein Binding; D013696:Temperature; D013997:Time Factors
chemical_list: D001672:Biocompatible Materials; D002234:Carbodiimides; D004798:Enzymes; D005656:Fungal Proteins; D003300:Copper; D048271:Chitosan; D042845:Laccase; D010100:Oxygen