Characterization and heterologous expression of laccase cDNAs from xylem tissues of yellow-poplar (Liriodendron tulipifera).
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 copper oxidases, 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.
Warnell School of Forest Resources, University of Georgia, Athens 30602, USA.
P R
PR
LaFayette
K E
KE
Eriksson
J F
JF
Dean
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D006031:Glycosylation; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics*; D042845:Laccase; D008031:Lignin / Q000302:isolation & purification; D019684:Magnoliopsida / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D014442:Monophenol Monooxygenase / Q000302:isolation & purification; D005810:Multigene Family; D010088:Oxidoreductases / Q000096:biosynthesis / Q000235:genetics*; D018547:Plant Stems / Q000201:enzymology / Q000235:genetics*; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution; D014026:Nicotiana / Q000235:genetics; D014197:Trees / Q000201:enzymology / Q000235:genetics*; D014934:Wood
D018076:DNA, Complementary; D007527:Isoenzymes; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D008031:Lignin; D010088:Oxidoreductases; D042845:Laccase; D014442:Monophenol Monooxygenase