Large-scale isolation, fractionation, and purification of soluble starch-synthesizing enzymes: starch synthase and branching enzyme from potato tubers.
Summary of "Large-scale isolation, fractionation, and purification of soluble starch-synthesizing enzymes: starch synthase and branching enzyme from potato tubers."
Soluble starch-synthesizing enzymes, starch synthase (SSS) and starch-branching enzyme (SBE), were isolated, fractionated, and purified from white potato tubers (Solanum tuberosum) on a large scale. Five steps were used: potato tuber extract from 2 kg of peeled potatoes, two acetone precipitations, and two fractionations on a large ultrafiltration polysulfone hollow fiber 100 kDa cartridge. Three kinds of fractions were obtained: (1) mixtures of SSS and SBE; (2) SSS, free of SBE; and (3) SBE, free of SSS. Contaminating enzymes (amylase, phosphorylase, and disproportionating enzyme) and carbohydrates were absent from the 2nd acetone precipitate and from the column fractions, as judged by the Molisch test and starch triiodide test. Activity yields of 122% (300,000-400,000 units) of SSS fractions and 187% (40,000-50,000 units) of SBE fractions were routinely obtained from the cartridge. Addition of 0.04% (w/v) polyvinyl alcohol 50K and 1 mM dithiothreitol to the glycine buffer (pH 8.4) gave long-term stability and higher yields of SSS and SBE, due to activation of inactive enzymes. Several SSS and SBE fractions from the two fractionations had very high specific activities, indicating high degrees of purification. Polyacrylamide gel electrophoresis of selected SSS and SBE fractions gave two to five SSS and/or SBE activity bands, corresponding to the one to five protein bands present in the 2nd acetone precipitate.
Laboratory of Carbohydrate Chemistry and Enzymology, Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011, USA.
This article was published in the following journal.
Name: Carbohydrate research
- PubMed Source: http://www.ncbi.nlm.nih.gov/pubmed/20620253
- DOI: http://dx.doi.org/10.1016/j.carres.2010.04.022
A method for purification of tRNA nucleotidyltransferase from Lupinus luteus L. seeds on a preparative scale is presented. This involved ammonium sulfate fractionation and chromatography on DEAE-cellu...
An extracellular, debranching isoamylase fromHendersonula toruloidea ATCC 64930, grown on starch, was purified 12-fold to an electrophoretically homogeneous state. The purified enzyme (estimated mol w...
Alcoholic-alkaline treatment of starch results in granular cold water-soluble starch. The nature of structural alterations occurred in starch due to the treatment is however relatively vague. Potato s...
A newly isolated, anaerobic, mesophillic bacterium, Clostridium sp. strain YK-3, ferments pentoses, hexoses, oligosaccharides and polysaccharides, such as soluble starch and glycogen, to ethanol and a...
Pancreatic islet transplantation is a promising therapy for Type I Diabetes. For many years the method used worldwide for islet purification in both rodent and human islet isolation has been Ficoll-ba...
This study will test the effects of resistant starch type 4 on blood sugar and hunger in adults with Type 2 diabetes.
To determine the effects of a commercially available dietary modification plus nutritional supplement regimen, the Standard Process 21 day Purification Program (SPPP), on serum lipids, C-r...
The purpose of this study is to determine the effects of a dietary fiber, resistant starch, on insulin sensitivity. Low insulin sensitivity is a risk factor for some diseases including ty...
The aim of our study was to evaluate the value of two different altered fractionation schedules – hyperfractionation and accelerated fractionation with concomitant boost – in compariso...
When people undergo major surgery, they require intravenous supplementation of fluids for a number of reasons: - to compensate for no oral intake - to support blood pressure...
Medical and Biotech [MESH] Definitions
The extent to which an enzyme retains its structural conformation or its activity when subjected to storage, isolation, and purification or various other physical or chemical manipulations, including proteolytic enzymes and heat.
A scale comprising 18 symptom constructs chosen to represent relatively independent dimensions of manifest psychopathology. The initial intended use was to provide more efficient assessment of treatment response in clinical psychopharmacology research; however, the scale was readily adapted to other uses. (From Hersen, M. and Bellack, A.S., Dictionary of Behavioral Assessment Techniques, p. 87)
Electrophoresis in which a starch gel (a mixture of amylose and amylopectin) is used as the diffusion medium.
The large scale production of pharmaceutically important and commercially valuable RECOMBINANT PROTEINS.
A scale that assesses the outcome of serious craniocerebral injuries, based on the level of regained social functioning.