%0 Journal Article %1 TPJ:TPJ12468 %A Ade, Carsten P %A Bemm, Felix %A Dickson, James M J %A Walter, Christian %A Harris, Philip J %D 2014 %J The Plant Journal %K Pinus cell galactoglucomannans glycosyltransferases loblolly pine plant radiata taeda walls xyloglucans %R 10.1111/tpj.12468 %T Family 34 glycosyltransferase (GT34) genes and proteins in Pinus radiata (radiata pine) and Pinus taeda (loblolly pine) %U http://dx.doi.org/10.1111/tpj.12468 %X Using a functional genomics approach, four candidate genes (PtGT34A, PtGT34B, PtGT34C and PtGT34D) were identified in Pinus taeda that encode CAZy family GT34 glycosyltransferases involved in the synthesis of cell-wall xyloglucans and heteromannans. The full-length coding sequences of three orthologs PrGT34A, B, and C, were isolated from a xylem-specific cDNA library from the closely related Pinus radiata. PrGT34B is the ortholog of XXT1 and XXT2, the two main xyloglucan (1→6)-α-xylosyltransferases in Arabidopsis thaliana. PrGT34C is the ortholog of XXT5 in A. thaliana, which is also considered to be involved in the xylosylation of xyloglucans. PrGT34A is an ortholog of a fenugreek galactosyltransferase involved in galactomannan synthesis. Truncated coding sequences of the genes were cloned into plasmid vectors and expressed in a Sf9 insect cell-culture system. The heterologous proteins were purified and in vitro assays showed that when incubated with UDP-xylose and cellotetraose, cellopentaose or cellohexaose, PrGT34B showed xylosyltransferase activity, and when incubated with UDP-galactose and the same cellooligosaccharides, showed some galactosyltransferase activity. The ratio of xylosyl- to galactosyltransferase activity was 434:1. Hydrolysis of the galactosyltransferase reaction products with galactosidases showed the linkages formed were α-. Analysis of the products of PrGT34B by matrix-assisted laser-desorption ionization time-of-flight mass spectrometry showed up to three xylosyl residues were transferred from UDP-xylose to cellohexaose. Heterologous proteins PrGT34A and PrGT34C showed no detectable enzymatic activity.This article is protected by copyright. All rights reserved.

@article{TPJ:TPJ12468, abstract = {Using a functional genomics approach, four candidate genes (PtGT34A, PtGT34B, PtGT34C and PtGT34D) were identified in Pinus taeda that encode CAZy family GT34 glycosyltransferases involved in the synthesis of cell-wall xyloglucans and heteromannans. The full-length coding sequences of three orthologs PrGT34A, B, and C, were isolated from a xylem-specific cDNA library from the closely related Pinus radiata. PrGT34B is the ortholog of XXT1 and XXT2, the two main xyloglucan (1→6)-α-xylosyltransferases in Arabidopsis thaliana. PrGT34C is the ortholog of XXT5 in A. thaliana, which is also considered to be involved in the xylosylation of xyloglucans. PrGT34A is an ortholog of a fenugreek galactosyltransferase involved in galactomannan synthesis. Truncated coding sequences of the genes were cloned into plasmid vectors and expressed in a Sf9 insect cell-culture system. The heterologous proteins were purified and in vitro assays showed that when incubated with UDP-xylose and cellotetraose, cellopentaose or cellohexaose, PrGT34B showed xylosyltransferase activity, and when incubated with UDP-galactose and the same cellooligosaccharides, showed some galactosyltransferase activity. The ratio of xylosyl- to galactosyltransferase activity was 434:1. Hydrolysis of the galactosyltransferase reaction products with galactosidases showed the linkages formed were α-. Analysis of the products of PrGT34B by matrix-assisted laser-desorption ionization time-of-flight mass spectrometry showed up to three xylosyl residues were transferred from UDP-xylose to cellohexaose. Heterologous proteins PrGT34A and PrGT34C showed no detectable enzymatic activity.This article is protected by copyright. All rights reserved.}, added-at = {2014-02-16T21:06:14.000+0100}, author = {Ade, Carsten P and Bemm, Felix and Dickson, James M J and Walter, Christian and Harris, Philip J}, biburl = {https://www.bibsonomy.org/bibtex/2e3f2deb3a6879f11578265c1ac5e176f/fbemm}, doi = {10.1111/tpj.12468}, interhash = {26672a08878a40141821a3e444f0d156}, intrahash = {e3f2deb3a6879f11578265c1ac5e176f}, issn = {1365-313X}, journal = {The Plant Journal}, keywords = {Pinus cell galactoglucomannans glycosyltransferases loblolly pine plant radiata taeda walls xyloglucans}, timestamp = {2014-02-16T21:06:14.000+0100}, title = {Family 34 glycosyltransferase (GT34) genes and proteins in Pinus radiata (radiata pine) and Pinus taeda (loblolly pine)}, url = {http://dx.doi.org/10.1111/tpj.12468}, year = 2014 }