Use of Dextran as a Ligand Carrier

Dextrans of the molecular weights normally used (40 000 and 500 000 Da) contain many thousands of reactive hydroxyl groups per molecule. The affinity partitioning effect achieved by introducing

Figure 10 (A) Scheme of the countercurrent distribution (CCD) process. (Reprinted from Johansson G, Andersson M and Akevland HE (1984) JournalofChromatography 298: 485-495. With permission from Elsevier Science.) (B) Distribution of protein and some glycolytic enzymes after CCD of an extract of bakers' yeast using 55 transfers. Without ligand-PEG (x); with Procion Olive MX-3G PEG, 1 % of total PEG (□); and with Procion yellow HE-3G PEG, 1 % of total PEG (O). System composition: 7% w/w dextran 500 and 5%w/w PEG 8000 including ligand-PEG, 50 mM sodium phosphate buffer pH 7.0, 0.2 mM EDTA, and 5 mM 2-mercaptoethanol. Temperature, 3°C. Systems in chamber 0-2 were initially loaded with yeast extract.

Figure 10 (A) Scheme of the countercurrent distribution (CCD) process. (Reprinted from Johansson G, Andersson M and Akevland HE (1984) JournalofChromatography 298: 485-495. With permission from Elsevier Science.) (B) Distribution of protein and some glycolytic enzymes after CCD of an extract of bakers' yeast using 55 transfers. Without ligand-PEG (x); with Procion Olive MX-3G PEG, 1 % of total PEG (□); and with Procion yellow HE-3G PEG, 1 % of total PEG (O). System composition: 7% w/w dextran 500 and 5%w/w PEG 8000 including ligand-PEG, 50 mM sodium phosphate buffer pH 7.0, 0.2 mM EDTA, and 5 mM 2-mercaptoethanol. Temperature, 3°C. Systems in chamber 0-2 were initially loaded with yeast extract.

Figure 11 (A) Partitioning of Procion yellow HE-3G dextran 70 (PrY-Dx) depending on the degree of substitution, n (expressed in molecules of dye bound per molecule of dextran), in systems containing 50 mM sodium phosphate buffer (A); 10 mM sodium sulfate (A); 100 mM sodium acetate (□); 100 mM KCl and 5 mM sodium phosphate buffer (•); or 100 mM KClO4 (O), at pH 7.9. Arrow indicates Kof unsubstituted dextran. System composition: 8%w/wdextran 70 and 4.5%w/w PEG 8000 including PrY-Dx (50 ^M bound dye), and indicated salt. Temperature, 22°C and pH of system adjusted to 7.9. (Reprinted from Johansson G and Joelsson M (1987) Journal ofChromatography 411: 161-166. With permission from Elsevier Science.) (B) Effect of the concentration of PrY-Dx on the partitioning of the enzyme glucose-6-phosphate dehydrogenase (G6PDH) using PrY-Dx with n equal to 1.3, □; 2.3, ■; 5.3, O; and 8.3, •. System as in (A) with 50 mM sodium phosphate buffer.

Figure 11 (A) Partitioning of Procion yellow HE-3G dextran 70 (PrY-Dx) depending on the degree of substitution, n (expressed in molecules of dye bound per molecule of dextran), in systems containing 50 mM sodium phosphate buffer (A); 10 mM sodium sulfate (A); 100 mM sodium acetate (□); 100 mM KCl and 5 mM sodium phosphate buffer (•); or 100 mM KClO4 (O), at pH 7.9. Arrow indicates Kof unsubstituted dextran. System composition: 8%w/wdextran 70 and 4.5%w/w PEG 8000 including PrY-Dx (50 ^M bound dye), and indicated salt. Temperature, 22°C and pH of system adjusted to 7.9. (Reprinted from Johansson G and Joelsson M (1987) Journal ofChromatography 411: 161-166. With permission from Elsevier Science.) (B) Effect of the concentration of PrY-Dx on the partitioning of the enzyme glucose-6-phosphate dehydrogenase (G6PDH) using PrY-Dx with n equal to 1.3, □; 2.3, ■; 5.3, O; and 8.3, •. System as in (A) with 50 mM sodium phosphate buffer.

one or just a few dye ligands is shown in Figure 11. Since the dye ligands used here carries seven to ten charged groups per molecule they also add a considerable (negative) net charge to the ligand dextran. Its partitioning will then be sensitive to the presence of salt and the choice of salt. The ligand-dextran can be directed either to the bottom phase or the top phase. This steering is more effective the greater the number of ligands per dextran molecule.

The effect of ligand-dextran on the partitioning of an enzyme, glucose-6-phosphate dehydrogenase, is shown in Figure 11. There is also a tendency towards affinity precipitation when the concentration of ligand molecules is equal to the concentration of enzyme binding sites in the system. This is seen as a shallow dip in the extraction curve.

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Solar Panel Basics

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