Characterization of two laccases of loblolly pine (Pinus taeda) expressed in tobacco BY-2 cells.
We previously showed that eight laccase genes (Lac 1-Lac 8) are preferentially expressed in differentiating xylem and are associated with lignification in loblolly pine (Pinus taeda) [Sato et al. (2001) J Plant Res 114:147-155]. In this study we generated transgenic tobacco suspension cell cultures that express the pine Lac 1 and Lac 2 proteins, and characterized the abilities of these proteins to oxidize monolignols. Lac 1 and Lac 2 enzymatic activities were detected only in the cell walls of transgenic tobacco cells, and could be extracted with high salt. The optimum pH for laccase activity with coniferyl alcohol as substrate was 5.0 for Lac 1 and between 5.0 and 6.0 for Lac 2. The activities of Lac 1 and Lac 2 increased as the concentration of CuSO(4) in the reaction mixtures increased in the range from 1 to 100 microM. Both enzymes were able to oxidize coniferyl alcohol and to produce dimers of coniferyl alcohol. These results are consistent with the hypothesis that Lac 1 and Lac 2 are involved in lignification in differentiating xylem of loblolly pine.
Sato
Yasushi
Y
Department of Biology, Graduate School of Science and Engineering, Ehime University, Matsuyama, Japan. ysato@sci.ehime-u.ac.jp
Whetten
Ross W
RW
D002461:Cell Line, Transformed; D003001:Cloning, Molecular; D019327:Copper Sulfate / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D042845:Laccase / Q000737:chemistry* / Q000235:genetics* / Q000502:physiology; D008031:Lignin / Q000378:metabolism; D010636:Phenols / Q000378:metabolism; D010666:Phenylpropionates / Q000378:metabolism; D041603:Pinus taeda / Q000201:enzymology*; D030821:Plants, Genetically Modified; D014026:Nicotiana; D052584:Xylem / Q000201:enzymology
D007527:Isoenzymes; D010636:Phenols; D010666:Phenylpropionates; C496130:sinapyl alcohol; D008031:Lignin; C010559:coniferyl alcohol; D042845:Laccase; D019327:Copper Sulfate