In this study, alpha-Galactosidase (alpha-D-galactoside-galactohydrolase, EC 3.2.1.22) has been isolated, purified, and characterized from intracellular fraction of Lactobacillus helveticus ATCC 10797. It was purified by a combination of ammonium sulfate precipitation and fast performance liquid chromatography system using ion exchange and gel-filtration columns. This enzyme was purified to only 9.44 fold over the crude extract with a recovery of 1.8%. The km of 3.83 mM and Vmax of 416.44 mumol/min/mg protein were calculated from PNPG. The molecular mass was estimated to be 188 kDa by gel-filtration, but 90 kDa by SDS-PAGE, indicating two similar molecular weight subunits. The optimum temperature for enzyme activity was 37°C, but with a stability below 30°C. The optimum pH was at 6 with a stability of pH 4-8 range. / This enzyme was activated by 10 mM monovalent ions such as K+ , NH4+, Li+ and CS +, while the activity was inhibited by divalent ions such as Cu +2, Zn+2, F+2. About 40% of the enzyme activity was inhibited with 100 mM EDTA. alpha-Galactosidase was inhibited by 1mM glucose and galactose, 10 mM sucrose, high concentrations of melibiose, or raffinose and stachyose, but the least inhibitory effect was shown with fructose. / When the sugars were incubated with alpha-galactosidase, melibiose was hydrolyzed to glucose and galactose, raffinose to galactose and sucrose, while stachyose to galactose and sucrose with raffinose as intermediate product.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.97889 |
| Date | January 2006 |
| Creators | Al-kandari, Sharifa. |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Master of Science (Department of Food Science and Agricultural Chemistry.) |
| Rights | © Sharifa Al-kandari, 2006 |
| Relation | alephsysno: 002482666, proquestno: AAIMR24598, Theses scanned by UMI/ProQuest. |
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