Expression and Characterization of Laccase Lac1 from Coriolopsis trogii Strain Mafic-2001 in Pichia pastoris and Its Degradation of Lignin

Abstract

The laccase gene (Lac1) was cloned from Coriolopsis trogii strain Mafic-2001. Full-length sequence of Lac1 containing 11 exons and 10 introns is composed of 2140 nucleotides (nts). mRNA of Lac1 encoded for a protein of 517 aa. Nucleotide sequence of the laccase was optimized and expressed in Pichia pastoris X-33. SDS-PAGE analysis showed that the molecular weight of the purified recombinant laccase rLac1 was about 70 kDa. The optimum temperature and pH of rLac1 were 40 ℃ and 3.0, respectively. rLac1 showed high residual activity (90%) in the solutions after 1 h incubation at the pH ranging from 2.5 to 8.0. rLac1 maintained over 60% of laccase activity at the temperatures ranging from 20 to 60 °C, and kept higher than 50% of its activity at 40 °C for 2 h. The activity of rLac1 was promoted by Cu²⁺ and inhibited by Fe²⁺. Under optimal conditions, lignin degradation rates of rLac1 on the substrates of rice straw, corn stover, and palm kernel cake were 50.24%, 55.49%, and 24.43% (the lignin contents of substrates untreated with rLac1 were 100%), respectively. Treated with rLac1, the structures of agricultural residues (rice straw, corn stover, and palm kernel cake) were obviously loosened which was reflected by the analysis of scanning electron microscopy and Fourier transform infrared spectroscopy. Based on the specific activity of rLac1 on the degradation of lignin, rLac1 from Coriolopsis trogii strain Mafic-2001 has the potential for in-depth utilization of agricultural residues.

Keywords: Coriolopsis trogii; Degradation; Laccase; Lignin,·Pichia pastoris.