Characterization and heterologous expression of laccase cDNAs from xylem tissues of yellow-poplar (Liriodendron tulipifera)

• Published in: Plant molecular biology Vol. 40; no. 1; pp. 23 - 35
• Main Authors: LaFayette, P.R, Eriksson, K.E.L, Dean, J.F.D
• Format: Journal Article
• Language: English
• Published: Netherlands Springer Nature B.V 01.05.1999
• Subjects: Amino Acid Sequence; amino acid sequences; Amino acids; Bacteria; binding sites; Cloning, Molecular; complementary DNA; Copper; DNA, Complementary - genetics; enzyme activity; genbank/af047697; genbank/u73103; genbank/u73104; genbank/u73105; genbank/u73106; Gene Expression; Glycosylation; histochemistry; isoelectric point; Isoenzymes - biosynthesis; Isoenzymes - genetics; isozymes; Laccase; Lignin - isolation & purification; Liriodendron tulipifera; Magnoliopsida - enzymology; Magnoliopsida - genetics; messenger RNA; Molecular Sequence Data; molecular weight; Monophenol Monooxygenase - isolation & purification; Multigene Family; Nicotiana - genetics; nucleotide sequences; Oxidoreductases - biosynthesis; Oxidoreductases - genetics; Plant Stems - enzymology; Plant Stems - genetics; Plants, Genetically Modified; Plants, Toxic; Protein Processing, Post-Translational; Protein Sorting Signals - genetics; Proteins; Recombinant Proteins - biosynthesis; Redox potential; Sequence Analysis, DNA; Sequence Homology, Amino Acid; Tissue Distribution; Trees - enzymology; Trees - genetics; Wood; xylem
• ISSN: 0167-4412; 1573-5028
• DOI: 10.1023/A:1026437406859

Four closely related cDNA clones encoding laccase isoenzymes from xylem tissues of yellow-poplar (Ltlacc2.1-4) were identified and sequenced. The inferred yellow-poplar laccase gene products were highly related to one another (79-91% at the amino acid level) and showed significant similarity to other blue copperoxidases, especially with respect to the copper-binding domains. The encoded proteins had N-terminal signal sequences and 17-19 potential N-linked glycosylation sites. The mature proteins were predicted to have molecular masses of ca. 61 kDa (unglycosylated) and high isoelectric points (pI 9.3-9.5). The canonical copper ligands were conserved, with the exception of a Leu residue associated with the axial position of the Type-1 cupric ion. The residue at this position has been proposed to influence the redox potential of Type-1 cupric ions. Northern blot analysis revealed that the yellow-poplar laccase genes are differentially expressed in xylem tissues. The genes were verified as encoding active laccases by heterologous expression in tobacco cells and demonstration of laccase activity in extracts from transformed tobacco cell lines.