Purification and characterization of stachyose synthase from lentil (Lens culinaris) seeds: galactopinitol and stachyose synthesis.
Stachyose synthase (STS) (EC 2.4.1.67) was purified 313-fold from mature seeds of lentil. The final preparation had a specific activity of 9.09 nkat stachyose formed per milligram of protein. The enzyme was a monomeric protein with a molecular mass of 88.6 kDa (SDS-PAGE) and an isoelectric point of 4.8 (chromatofocusing). Western analysis revealed cross-reactivity of polyclonal antibodies raised against STS from adzuki bean with the lentil enzyme. The purified enzyme catalyzed a range of different galactosyl transfer reactions. In addition to the genuine STS reaction (raffinose + galactinol --> stachyose + myo-inositol), the enzyme catalyzed the reversible galactosyl transfer from galactinol to d-pinitol (1d-3-O-methyl-chiro-inositol), yielding galactopinitol A (O-alpha-d-galactopyranosyl-(1 --> 2)-4-O-methyl-d-chiro-inositol) and myo-inositol. Galactopinitol A could be further galactosylated by STS to give ciceritol (O-alpha-d-galactopyranosyl-(1 --> 6)-O-alpha-d-galactopyranosyl-(1 --> 2)-4-O-methyl-d-chiro-inositol). Enzymatic synthesis of galactopinitol A and ciceritol is a new observation. However, STS was not only able to utilize galactopinitol A as galactosyl acceptor, but also as galactosyl donor to form stachyose from raffinose. The role of STS in the metabolism of galactosyl cyclitols and oligosaccharides in plant seeds is discussed.
Hoch
G
G
Institute of Plant Physiology, University of Vienna, Vienna, A-1091, Austria.
Peterbauer
T
T
Richter
A
A
D004187:Disaccharides / Q000096:biosynthesis* / Q000378:metabolism; D007887:Fabaceae / Q000201:enzymology*; D005700:Galactosyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D007294:Inositol / Q000378:metabolism; D007700:Kinetics; D009844:Oligosaccharides / Q000096:biosynthesis*; D010946:Plants, Medicinal; D011887:Raffinose / Q000378:metabolism; D012639:Seeds / Q000201:enzymology; D013379:Substrate Specificity
D004187:Disaccharides; D009844:Oligosaccharides; C119541:galactopinitol A; C005695:stachyose; C013536:6 beta-galactinol; D007294:Inositol; D005700:Galactosyltransferases; C113913:galactinol -raffinose galactosyltransferase; D011887:Raffinose