Processes for the purification of human recombinant decorin and

Chemistry: natural resins or derivatives; peptides or proteins; – Proteins – i.e. – more than 100 amino acid residues – Glycoprotein – e.g. – mucins – proteoglycans – etc.

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530412, 530416, C07K 120, C07K 1400

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active

055678076

ABSTRACT:
The present invention is directed to a process for the production of substantially pure human recombinant decorin which involves the combination of three separate stages characterized by contacting decorin-containing cell culture medium with (1) a first strong anionic exchange resin; then with (2) a hydrophobic interactive chromatographic resin; and finally with (3) a second strong anionic exchange resin. By using a combination of steps, and certain reagents, in particular a 2.4 to 3 molar GuHCl solution to elute decorin from a hydrophobic interactive column, the process of this invention provides a more convenient and reproducible process for purifying human recombinant decorin. The invention also provides a process for detecting the presence of guanidinium ions in a sample solution. The detection process involves contacting a sample solution suspected of containing guanidinium ions with a cation exchange resin and eluting the guanidinium ions present in tire sample solution with an aqueous buffer solution having a pH of about 1.5 to about 2. This is followed by contacting the eluant with a cation suppressor columns and simultaneously flowing a suppressor regenerate solution in the opposite direction on the opposite side of the permeable membrane of the column, and finally, detecting the presence of guanidinium ions in the eluant from the ion exchange column which was contacted with the suppressor column by use of a conductivity detector.

REFERENCES:
Takeuchi et al., J. Biol. Chem., 269(51); 32634-32638 (1994).
Vilim et al., Biochem. J., 304(3); 887-894 (1994).
Evanko et al., Arch. Biochem. Biophys. 307(1); 153-164 (1993).
Krusius and Ruoslahti, "Primary structure of an extracellular matrix proteoglycna core protein deduced from cloned cDNA." Proc. Natl. Acad. Sci. USA, 83:7683-7687 (1986).
Ruoslahti Erkki, "Structure and biology of proteoglycans." Ann. Rev. Cell. Bio., 4:229-255 (1988).
Choi et al., "Characterization of the dermatan sulfate proteoglycans, DS-PGI and DS-PGII, from bovine articular cartilage and skin isolated by octyl-sepharose chromatography." J. of Biol. Chem., 264(5):2876-2884 (1989).

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