title: Purification and characterization of a temperature- and pH-stable laccase from the spores of Bacillus vallismortis fmb-103 and its application in the degradation of malachite green.
abstract: Malachite green residue can affect aquaculture food safety. Bioremediation of contaminated water by enzyme treatment is an environmentally friendly and economical way to remove contaminating substances. In the present study, a temperature- and pH-stable laccase was purified from the spores of Bacillus ballismortis fmb-103 and was used to degrade malachite green. The laccase from fmb-103 (fmb-L103) was purified 15.2-fold to homogeneity (389.9 mU/mg protein with respect to ABTS as a substrate) by precipitation with 30-80% (NH4)2SO4, DEAE-Sephadex A-50 ion exchange chromatography, and Sephadex G-100 chromatography. fmb-L103 is a nonblue laccase with a molecular weight of 55.0 kDa and Cu content of 2.5 (mol:mol). fmb-L103 retained more than 50% activity after 10 h at 70 °C and demonstrated broad pH stability in both acidic and alkaline conditions. The effects of inhibitors and metal ions on fmb-L103 activity were also examined. A kinetic study revealed that ABTS was a suitable substrate with a Km of 22.7 μmol and a Vmax of 3.32 μmol/mL/min. fmb-L103 can efficiently degrade malachite green after a 48 h treatment period in combination with a mediator, without the appearance of leucomalachite green.
authors.affiliation: Laboratory of Enzyme Engineering, College of Food Science and Technology, Nanjing Agriculture University, Nanjing 210095, PR China.
authors.forename: Chong
authors.initials: C
authors.lastname: Zhang
authors.forename: Shuang
authors.initials: S
authors.lastname: Zhang
authors.forename: Hanwen
authors.initials: H
authors.lastname: Diao
authors.forename: Haizhen
authors.initials: H
authors.lastname: Zhao
authors.forename: Xiaoyu
authors.initials: X
authors.lastname: Zhu
authors.forename: Fengxia
authors.initials: F
authors.lastname: Lu
authors.forename: Zhaoxin
authors.initials: Z
authors.lastname: Lu
mesh_terms: D001407:Bacillus / Q000737:chemistry* / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D001673:Biodegradation, Environmental; D052918:Environmental Restoration and Remediation / Q000379:methods*; D004795:Enzyme Stability; D007700:Kinetics; D042845:Laccase / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D012394:Rosaniline Dyes / Q000737:chemistry*; D013172:Spores, Fungal / Q000737:chemistry / Q000201:enzymology*; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D012394:Rosaniline Dyes; C005095:malachite green; D042845:Laccase