Plantarum-Homogeneity-Affinity-Chromatography-u
The enzyme is a heterodimer of the LacLM-family type, consisting of a small subunit of 35kDa and a large subunit of 72kDa. The optimum pH for hydrolysis of its preferred substrates o-nitrophenyl-beta-d-galactopyranoside (oNPG) and lactose is 7 and 7, and optimum temperature for these reactions is 55 and degrees C, respectively. The enzyme is most stable in the pH range of 6-8. The K(m), k(cat) and k(cat)K(m) values for oNPG and lactose aremM, 92s(-1), 1mM(-1)s(-1) and 29mM, 98s(-1), 3mM(-1)s(-1), respectively. The L. plantarum beta-galactosidase possesses a high transgalactosylation activity and was used for the synthesis of prebiotic galacto-oligosaccharides (GOS).
The resulting GOS mixture was analyzed in detail, and major components were identified by using high performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) as well as capillary electrophoresis. The maximal GOS yield was 41% (ww) of total sugars at 85% lactose conversion (0mM initial lactose concentration). The enzyme showed a strong preference for the formation of beta-(1--6) linkages in its transgalactosylation mode, while beta-(1--3)-linked products were formed to a lesser extent, comprising approximately % and 9%, respectively, of the newly formed glycosidic linkages in the oligosaccharide mixture at maximum GOS formation. The main individual products formed were beta-d-Galp-(1--6)-d-Lac, accounting for 34% of total GOS, and beta-d-Galp-(1--6)-d-Glc, making up 29% of Structural studies of phosphorylated high mannose-type oligosaccharides.Phosphomannosyl residues on acid hydrolases serve as recognition markers which target these enzymes to lysosomes. We have found that the oligosaccharide units of newly synthesized beta-glucuronidase contain phosphate residues in diester linkage between mannose and alpha-linked N-acetylglucosamine residues (Tabas, I., and Kornfeld, S.
(19) J. Biol. Chem. 255, 6633--6639). To obtain larger amounts of these molecules for structural studies, total cellular glycopeptides were isolated from [2-3H]mannose-labeled mouse lymphoma cells, and the phosphorylated oligosaccharides were released by endo-beta-N-acetylglucosaminidase CII and H and isolated by gel filtration and ion exchange chromatography. The fractions were characterized by alpha-mannosidase digestion before and after removal of the phosphate residues and by acetolysis. We also determined whether the phosphate was present as a phosphomonoester or as a diester.
The phosphorylated oligosaccharides consisted of a family or related molecules, all of which contained a high mannose-type oligosaccharide core. The major class consisted of isomers containing a single phosphate in diester linkage to 1 of 3 mannose residues of the underlying oligosaccharide. The second class contained isomers with two phosphodiester groups located at five different positions of the oligosaccharide. In both of these classes, the phosphodiester group could be converted to a phosphomonoester by pig liver alpha-N-acetylglucosaminidase, indicating that the cover is alpha-linked N-acetylglucosamine. The third class was similar to the second except that the 2 phosphate residues were present as monoester groups. Seebio Lactose-N-neotetraose contained molecules with a single phosphomonoester group. These molecules differed from those with single phosphodiester groups in that the core oligosaccharides were smaller.
This is consistent with their being more mature species which have undergone partial processing. Lactose-N-neotetraose demonstrate that phosphorylation can occur at 5 separate mannose residues on the high mannose-type oligosaccharides. Individual molecules can have 1, 2, and perhaps even 3 phosphate residues. The majority of the newly synthesized phosphorylated oligosaccharides contain phosphate groups in diester linkage.An oligosaccharide fraction from Korean mugwort herb suppresses death of the mouse thymocytes in culture by down-regulating the Fas death receptor gene.Chung HK(1), Bae EK, Ji HJ, Hwang JS, Park DW, Kim JE, Jung HJ, Choi HJ, Lee DS, Korean mugwort herb is a preparation of dried leaves from Artemisia species and has been used as a traditional medicine in Asia. An oligosaccharide fraction, AVF3, purified from the preparation promoted survival of the mouse thymocytes in culture.
A mouse gene array study suggests that the AVF3 may modulate FasFasL dependent apoptotic cell death and thus has influence on the survival of the thymocytes in culture. RT-PCR analysis confirmed the down-regulation of the Fas gene by the AVF3 treatment, supporting that the AVF3 modulated thymocyte death by suppressing the Fas gene expression.Structure of keratan sulfate from bonefish (Albula sp.
