Method of separating and purifying two active forms of urokinase

Details Method of separating and purifying two active forms of urokinase Method of separating and purifying two active forms of urokinase

1976-10-08

1978-01-03

Jones, Raymond N.

Textiles: fiber preparation

195 62, C07G 702, C12D 1310

Patent

active

040665062

ABSTRACT:
An affinity chromatography method for the separation and pruification of two active forms of urokinase from a crude urokinase preparation. The method employs an extracting agent of agmatine covalently coupled to the surface of a water-insoluble solid support material such as agarose, and three separate specific buffer solutions. A low ionic strength buffer, such as 0.01 molar sodium phosphate buffer, pH 6.0-9.0, is used in preparing the loading solution and for washing the extracting agent. A slightly higher ionic strength buffer, such as 0.02 molar sodium phosphate buffer, pH 6.0-9.0, is used as a first eluant for eluting from the extracting agent a first active form of urokinase which is characterized by a molecular weight of approximately 33,400 and a specific activity of approximately 226,000 CTA units/mg protein. A still higher ionic strength buffer with added salt, such as 0.1 molar sodium phosphate -- 0.4 molar sodium chloride buffer, pH 5.0-8.0, is used as a second eluant for eluting from the extracting agent a second active form of urokinase which is characterized by a molecular weight of approximately 47,000 and a specific activity of approximately 104,000 CTA units/mg protein.

REFERENCES:
patent: 3355361 (1967-11-01), Lesuk
patent: 3819605 (1974-06-01), Holleman et al.
patent: 3879369 (1975-04-01), Nolan
Maciag et al., "Purification of Urokinase by Affinity Chromatography," Chemical Abstracts, vol. 83, No. 5, p. 183 (1975), Abs. No. 39272d.
Shiba et al., "Separation and Measurement of Plasminogen Activators from Tissue and Body Fluid by Affinity Chromatography.II. Separation of Urokinase from Human Urine," Chemical Abstracts, vol. 80, No. 11, p. 114, Abs. No. 56897s.
Holmberg et al., "Purification of Urokinase by Affinity Chromatography," Chemical Abstracts, vol. 85, No. 13, p. 205, (1976), Abs. No. 89077f.
Tamura et al., "Purification of Human Plasma Kallikrein and Urokinase by Affinity Chromatography," Chemical Abstracts, vol. 85, No. 23, p. 179 (1976), Abs. No. 173251r.
Holmberg et al., "Purification of Urokinase by Affinity Chromatography," Biochimica et Biophysica Acta, vol. 445, No. 1, (1976), pp. 215-222.

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