Article Figures & Data
Figures
- Fig. 1.
α-Galactosidase-catalyzed release of labeled Gal from the purified product of incubating mannoheptaose (M7) with UDP-(14C)Gal in the presence of digitonin-solubilized fenugreek GMGT. The α-galactosidase was diluted to give a slow reaction. Samples of the α-galactosidase reaction mixture were removed at various times and subjected to TLC. The figure shows a digital autoradiogram of the TLC plate. M7prod, Purified transfer product.
- Fig. 2.
β-Mannosidase digestion of a purified sample of labeled monogalactosylmannohexaose (M6G) formed by incubating mannohexaose with UDP-(14C)Gal in presence of Triton X-100-solubilized fenugreek GMGT. Samples of the β-mannosidase reaction mixture were taken at various times and subjected to TLC. The figure is a digital autoradiogram of the TLC plate. R, Reference standards of (14C)Man-labeled galactomannan oligosaccharides from endo-β-mannanase digestion of galactomannan biosynthesized in vitro (Reid et al., 1995).
- Fig. 3.
Observed pattern of Gal substitution of M5, M6, M7, and M8 by detergent-solubilized fenugreek GMGT in relation to a hypothetical GMGT acceptor substrate subsite recognition sequence comprising six Man residues (numbered 1–6 from the nonreducing end of the sequence), with transfer occurring to the Man residue at site 3 of the recognition sequence. An asterisk indicates an observed position of minor substitution. A double asterisk indicates an observed position of major substitution.
- Fig. 4.
A model for the interaction of MS and GMGT during galactomannan biosynthesis in fenugreek.
Tables
- Table 1.
Products of snail exo-β-d-mannosidase action on manno- and galactomanno-oligosaccharides
All mannosyl linkages are (1→4)-β, and galactosyl linkages are (1→6)-α. M, Mannosyl residue. G, Galactosyl residue.
- Table II.
Deduction and quantitative estimation of structural isomers comprising M5G, M6G, M7G, and M8G formed in experiments using digitonin- and Triton X-100-solubilized fenugreek GMGT
Compounds Surviving β-Mannosidase Action Deduced Structure before β-Mannosidase Treatment Relative Amount % Digitonin TX-100 se M5G MGMMM MGMMM 20 ± 3 26 ± 2 MGMM MMGMM 80 ± 3 75 ± 2 M6G MGMMMM MGMMMM 10 ± 3 5 ± 5 MGMMM MMGMMM 75 ± 4 83 ± 4 MGMM MMMGMM 16 ± 0 12 ± 2 M7G MGMMMMM MGMMMMM 8 ± 4 3 ± 3 MGMMMM MMGMMMM 41 ± 2 44 ± 4 MGMMM MMMGMMM 43 ± 2 44 ± 3 MGMM MMMMGMM 10 ± 2 9 ± 1 M8G MGMMMMMM MGMMMMMM 1 ± 1 32-a MGMMMMM MMGMMMMM 20 ± 4 202-a MGMMMM MMMGMMMM 34 ± 3 352-a MGMMM MMMMGMMM 30 ± 0 272-a MGMM MMMMMGMM 16 ± 5 152-a M, Unsubstituted mannose residue; G, galactose-substituted mannose residue; TX-100, Triton X-100. Structures read from nonreducing to reducing end. Thus,
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↵F2-a Single determination; otherwise, means of at least two independent experiments.
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- Table III.
Predicted and experimentally observed products of endo-β-mannanase digestion of labeled M5G, M6G, M7G and M8G formed using digitonin-solubilized fenugreek GMGT.
Deduced Structures of Positional Isomers with Predicted Labeled Endo-β-Mannanase Products Relative Proportions of Labeled M2G and M3G Released on Endo-β-Mannanase Digestion Predicted Experimental3-b M2G M3G M2G M3G M5G 20 ± 3 80 ± 3 24 ± 2 76 ± 2 MGMMM → M2G MMGMM → M3G M6G 26 ± 3 75 ± 4 25 ± 1 75 ± 1 MGMMMM → M2G MMGMMM → M3G MMMGMM → M2G M7G 61 ± 4 41 ± 2 57 ± 2 43 ± 2 MGMMMMM → M2G MMGMMMM → M3G MMMGMMM → M2G MMMMGMM → M2G M8G 73 ± 4 28 ± 4 68 ± 1 32 ± 1 MGMMMMMM → M2G MMGMMMMM → M3G MMMGMMMM → M2G MMMMGMMM → M2G MMMMMGMM → M2G +M3G3-b Experimental data were obtained by digesting M5G to M8G completely with A. niger endo-β-mannanase, separating the digests by TLC and determining the relative proportions of the product oligosaccharides M2G and M3G by quantitative digital autoradiography. Predictions were made using the known specificity of the endo-β-mannanase (McCleary and Matheson, 1983; summarized in text) and the digitonin data from Table II.
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F3-a Mean (± se) of at least two independent experiments.
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↵F3-b Assumed 50% of each.
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