Cloning and Expression Analysis of Vv-lac3, A Novel Functional Laccase Gene Involved in Stipe Elongation, Based on the Genomic Sequence of Volvariella volvacea
- *Corresponding Author:
- Baogui Xie
Mycological Research Center, College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, PR China
E-mail: [email protected]
Received date: 23 June 2015; Accepted date: 01 August 2015; Published date: 07 August 2015
Fungal laccases always play important roles in various physiological and developmental processes. Here, we cloned a laccase gene, and named Vv-lac3, cDNA from the Volvariella volvacea, and expressed the cDNA in Pichia pastoris. The Vv-lac3 cDNA consisted of 1599 bp containing an open reading frame (ORF) encoding a 532 amino acids. The analysis of the deduced amino acids revealed that Vv-lac3 possessed a 19-amino acids signal peptide at the N-terminal end and a 513-amino acid mature protein. The Vv-lac3 contained four conserved copper-binding sites that which is the typical structure of fungal laccases. Phylogenetic analysis showed that Vv-lac3 had a high degree of identity with other basidiomycete laccases. Native-PAGE and SDS-PAGE analysis demonstrated that the product of Vvlac3 cDNA from P. pastoris was a functional laccase with a molecular mass of 65 kDa. The expression of the Vv-lac3 gene in V. volvacea increased during button stage to the elongation stage; it reached peaked in the elongation stage, and then decreased in the maturation stage, which suggests that this gene plays a regulatory role in stipe elongation in V. volvacea.