Abstract
Pectin is one of the major cell wall polysaccharides found in dicotyledonous plants. We have solubilized and partially purified a β-(1→4)-galactosyltransferase (GalT) involved in the synthesis of the β-(1→4)-galactan side chains of pectin. The enzyme protein was almost completely solubilized by mixing a crude microsomal preparation of etiolated 6-day-old soybean (Glycine max Merr.) hypocotyls with a detergent, Triton X-100 (0.75%, w/v), in buffer. The solubilized enzyme was partially purified by ion-exchange chromatography. The crude membrane-bound GalT transferred Gal from UDP-Gal onto 2-aminobenzamide (AB)-derivatized β-(1→4)-galactoheptaose (Gal7-AB), leading to the formation of Gal8–11-AB by attachment of a series of one to four galactosyl residues; this is similar to what has previously been observed for 2-aminopyridine-derivatized β-(1→4)-galactooligomer acceptors (Konishi et al. in Planta 218:833–842, 2004). The partially purified GalT, by contrast, was able to transfer more than 25 galactosyl residues and elongated the chains to about Gal35-AB, thus almost reaching the length (43–47 Gal units) of native β-(1→4)-galactan side chains found in pectic polysaccharides from soybean cotyledons (Nakamura et al. in Biosci Biotechnol Biochem 66:1301–1313, 2002). Enzyme activity increased with increasing chain length of β-(1→4)-galactooligomers and reached maximal activity at heptaose, whereas galactooligomers higher than heptaose showed lower acceptor efficiency.
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Abbreviations
- AB:
-
2-Aminobenzamide
- DP:
-
Degree of polymerization
- GalA:
-
Galacturonic acid
- GalT:
-
Galactosyltransferase
- IMO:
-
Isomaltooligomer
- MALDI-TOF/MS:
-
Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry
- RG:
-
Rhamnogalacturonan
- l-Rha:
-
l-Rhamnose
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Acknowledgment
We thank Dr. H. Nakano, Osaka Municipal Research Institute, for the kind gift of endo-β-(1→4)-galactanase.
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Sugars described in this paper belong to the d-series unless otherwise noted.
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Konishi, T., Kotake, T. & Tsumuraya, Y. Chain elongation of pectic β-(1→4)-galactan by a partially purified galactosyltransferase from soybean (Glycine max Merr.) hypocotyls. Planta 226, 571–579 (2007). https://doi.org/10.1007/s00425-007-0505-3
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DOI: https://doi.org/10.1007/s00425-007-0505-3