First published online March 19, 2004; 10.1104/pp.103.036210
Plant Physiology 134:1377-1387 (2004)
© 2004 American Society of Plant Biologists
BIOCHEMICAL PROCESSES AND MACROMOLECULAR STRUCTURES
Cloning, Functional Expression, and Characterization of the Raffinose Oligosaccharide Chain Elongation Enzyme, Galactan:Galactan Galactosyltransferase, from Common Bugle Leaves1
Esther M. Tapernoux-Lüthi,
Andreas Böhm and
Felix Keller*
Institute of Plant Biology, University of Zurich, Zollikerstrasse 107, CH–8008 Zurich, Switzerland
Galactan:galactan galactosyltransferase (GGT) is a unique enzyme of the raffinose family oligosaccharide (RFO) biosynthetic pathway. It catalyzes the chain elongation of RFOs without using galactinol ( -galactosyl-myoinositol) by simply transferring a terminal -galactosyl residue from one RFO molecule to another one. Here, we report the cloning and functional expression of a cDNA encoding GGT from leaves of the common bugle (Ajuga reptans), a winter-hardy long-chain RFO-storing Lamiaceae. The cDNA comprises an open reading frame of 1215 bp. Expression in tobacco (Nicotiana plumbaginifolia) protoplasts resulted in a functional recombinant protein, which showed GGT activity like the previously described purified, native GGT enzyme. At the amino acid level, GGT shows high homologies (>60%) to acid plant -galactosidases of the family 27 of glycosylhydrolases. It is clearly distinct from the family 36 of glycosylhydrolases, which harbor galactinol-dependent raffinose and stachyose synthases as well as alkaline -galactosidases. Physiological studies on the role of GGT confirmed that GGT plays a key role in RFO chain elongation and carbon storage. When excised leaves were exposed to chilling temperatures, levels of GGT transcripts, enzyme activities, and long-chain RFO concentrations increased concomitantly. On a whole-plant level, chilling temperatures induced GGT expression mainly in the roots and fully developed leaves, both known RFO storage organs of the common bugle, indicating an adaptation of the metabolism from active growth to transient storage in the cold.
1 This work was supported by the Swiss National Foundation.
Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/doi/10.1104/pp.103.036210.
* Corresponding author; e-mail fkel{at}botinst.unizh.ch; fax +41 1 634 82 04.
Received November 14, 2003;
returned for revision December 23, 2003;
accepted January 19, 2004.
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