Abstract
Studies of the interactions of Cibacron Blue F3GA in aqueous solutions with organic solvents, salts, oligo-, and poly-peptides by visible absorption difference spectroscopy indicate that the dye-sodium chloride (or any other salt) interaction results in a difference spectrum that has a characteristic positive peak at 690 nm and negative double minima at 630 and 585 nm. Such a “salt-like” or “ionic” spectrum is also obtained with polycations such as spermine, spermidine, oligolysines, polylysine, polyarginine, and protamine. In a striking contrast, the difference spectrum of the dye in binary aqueous solvents containing dioxan or t-butyl alcohol at moderately high concentrations, measured against water, displays a positive peak and shoulder at 655 and 610 nm, respectively with a small negative contribution at wavelengths less than 550 nm. We assign this type of spectrum to a “nonpolar” interaction of the dye with the organic cosolvent molecules. Such spectral characteristics have been widely noticed in several studies involving Cibacron Blue and proteins, thereby providing a basis for a reasonable interpretation for the interaction of the dye with the proteins. An understanding of the forces involved in dye-protein interactions is essential for protein purification studies using the technique of dye-ligand chromatography. Complete separation of pepsin and chymosin present in calf rennet has been achieved using Blue agarose chromatography. The separation appears to result from a combination of hydrophobic and electrostatic interactions of chymosin with the dye ligand. Differential surface hydrophobicity of the two enzymes may also play a key role in the separation.
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References
Ryan, L.D., and Vestling, C.S., Rapid purification of lactate dehydrogenase from rat liver and hepatoma a new approach. Arch. Biochem. Biophys. 1974, 160, 279–284
Bohme, H.J., Kopperschlager, G., Schulz., and Hoffman E., Affinity chromatography of phosphofructokinase using Cibacron Blue F3GA. J. Chromatogr., 1972, 69, 209–214
Stellwagen E., Use of Blue Dextran as a probe for the nicotinamide adenine dinucleotide domain in proteins. Acc. Chem. Res., 1977, 10, 92–98
Thompson S.T., and Stellwagen E., Binding of Cibacron Blue F3GA to proteins containing the dinucleotide fold. Proc. Natl. Acad. Sci. U.S.A., 1976, 73, 361–365
Kaufman B.T., and Kemerer V.F., Characterization of chicken liver dihydrofolate reductase after purification by affinity chromatography and isoelectric focusing. Arch. Biochem. Biophys., 1977, 179, 420–431
International Dairy Federation Standard, 110 1982
Subramanian S., Separation of chymosin and pepsin in calf rennet by dyeligand affinity chromatography. Prep. Biochem. 1987, 17, 297–312
Edwards R.A. and Woody R.W., Induced circular dichroism as a probe of Cibacron Blue and Congo Red bound to dehydrogenases. Biochem. Biophys. Res. Commun., 1977, 79, 470–476
Subramanian S., Specific interaction of spermine with Cibacron Blue F3GA. Arch. Biochem. Biophys., 1982, 217, 388–391
Subramanian S., Spectral changes induced in Cibacron Blue F3GA by salts, organic solvents, and polypeptides; Implications for blue dye interaction with proteins. Arch. Biochem. Biophys., 1982, 216, 116–125
Pompon D., and Lederer F., Binding of Cibacron Blue F3GA to Flavocytochrome b2 from Baker’s yeast. Eur. J. Biochem. 1978, 90, 563–569
Biellmann J-F., Samama J-P., Branden C-I., and Eklund H., X-ray studies of the binding of Cibacron Blue F3GA to liver alcohol dehydrogenase. Eur. J. Biochem., 1979, 102, 107–110
Barden R.E., Darke P.L., Deems R.A., and Dennis E.A., Interaction of phospholipase A2 from cobra venom with Cibacron Blue F3GA. Biochemistry, 1980, 19, 1621–1625
Subramanian S., and Kaufman B.T., Dihydrofolate reductase from chicken liver and Lactobacillus casei bind Cibacron Blue F3GA in different modes and at different sites. J. Biol. Chem. 1980, 225, 10587–10590
Appukuttan P.S., and Bachhawat B.K., Separation of polypeptide chains of ricin and the interaction of the A chain with Cibacron Blue F3GA. Biochem Biophys. Acta, 1979, 580, 10–14
Bull P., MacDonald H., and Valenzuela P., The interaction of yeast RNA polymerase I and Cibacron Blue F3GA. Biochim. Biophys. Acta, 1981, 653, 368–377
Pompon D., Guiard B. and Lederer F., Binding of Cibacron Blue F3GA to the flavin and NADH sites in cytochrome b5 reductase. Eur. J. Biochem. 1980, 110, 565–570
Jankowski W.J., von Muenchhausen W., Sulkowski E. and Carter W.A., Binding of human interferons to immobilized Cibacron Blue F3GA the nature of molecular interaction. Biochemistry, 1976, 15, 5182–5187
Foltmann B., Prochymosin and chymosin (Prorennin and Rennin) in Methods in Enzymology, Vol. XIX, Ed. G.E. Perlmann and L. Lorand, 1970, pp 421–436
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© 1989 Elsevier Science Publishers Ltd
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Subramanian, S. (1989). Ionic and Apolar Interactions of Cibacron Blue F3GA with Model Compounds and Proteins Application to Protein Purification. In: Vijayalakshmi, M.A., Bertrand, O. (eds) Protein-Dye Interactions: Developments and Applications. Springer, Dordrecht. https://doi.org/10.1007/978-94-009-1107-9_6
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DOI: https://doi.org/10.1007/978-94-009-1107-9_6
Publisher Name: Springer, Dordrecht
Print ISBN: 978-94-010-6989-2
Online ISBN: 978-94-009-1107-9
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