Method for highly purifying human serum albumin

Chemistry: natural resins or derivatives; peptides or proteins; – Proteins – i.e. – more than 100 amino acid residues – Albumin

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530413, C12P 2100

Patent

active

056567294

DESCRIPTION:

BRIEF SUMMARY
TECHNICAL FIELD

The present invention relates to a method for highly purifying human serum albumin, which is characterized by subjecting human serum albumin produced by genetic engineering to Cu-chelating chromatography.


BACKGROUND ART

An albumin, particularly human serum albumin (hereinafter also referred to as HSA) is an important component constituting protein in plasma. This protein is produced in liver, and is mainly responsible for sustaining normal osmotic pressure of blood flow. Also, it functions as a carrier for various serum molecules. HSA is administered in a variety of clinical situations. For example, when HSA is administered to a patient suffering from shock or ambustion, it functions to recover blood volume to its original level, thereby improving some symptoms relating to trauma. For this effect, HSA is frequently administered. Also, patients suffering from hypoproteinemia or fetal erythroblastosis may need treatments with HSA.
As exemplified, the basic significance of HSA administration is prominent in the treatment of symptoms accompanying loss of fluids from blood vessels, as in surgery, shock, burn, or hypoproteinemia which causes edema.
At present, HSA is produced mainly by fractionation of blood. This production method is uneconomical, and besides, it poses a problem that the supply of the blood is not always assured. Moreover, the blood may contain undesirable substances such as hepatitis viruses. Accordingly, the development of a substitute raw material for HSA will be greatly advantageous.
In the meantime, the advent of the recombinant DNA technique has enabled production of various useful polypeptides by microorganisms, and many mammalian polypeptides have been already produced by various kinds of microorganisms. A technique permitting large-scale production of HSA by utilizing genetic engineering and purification thereof is being established.
Methods for isolating and purifying HSA from plasma have been variously studied and have seen practical application. For example, Cohn's ethanol fractionation, PEG fractionation and ammonium sulfate fractionation are known. In recent years, a combined method of treatment with anion exchanger and heat treatment at 60.degree. C. for 10.hours (Japanese Patent Unexamined Publication No. 191226/1990), and a combined method of treatment with anion exchanger, treatment with cation exchanger and heat treatment at 60.degree. C. for 10 hours (Japanese Patent Unexamined Publication No. 17123/1991) have been developed.
While the purification of recombinant HSA (r-HSA) obtained by genetic engineering has been studied in a variety of ways, the study has not yet succeeded in removing contaminant components derived from yeast. The presence of such component derived from yeast has a probability of causing problems of antigenicity, since the component is a foreign substance to living organisms. That is, the purity of the recombinant type proteins is insufficient and the contaminant component derived from yeast needs to be removed further.


DISCLOSURE OF THE INVENTION

An object of the present invention is to remove, when producing HSA by genetic engineering, the aforementioned component derived from yeast, which cannot be sufficiently removed by conventional methods for purifying HSA produced by genetic engineering, and to provide a highly purified HSA.
In view of the above situation, the present inventors conducted various studies for achieving the object as described, and have found that Cu-chelating chromatography of HSA for purification thereof, when obtaining HSA by genetic engineering, results in advanced removal of the contaminant components derived from yeast, which resulted in the completion of the invention.
Accordingly, the present invention relates to a method for highly purifying human serum albumin, which comprises subjecting a fraction containing human serum albumin produced by genetic engineering, to Cu-chelating chromatography, and more specifically, the present invention relates to a method for highly purifying human serum albumin prod

REFERENCES:
patent: 5169936 (1992-12-01), Staples et al.
patent: 5330901 (1994-07-01), Prevatt et al.
patent: 5440018 (1995-08-01), Ohmura et al.
Hansson et al., J. Chromatog. 215:333-339 (1981).
Sulkowski, Trends Biotechnol. 3:1-7 (1985).
Andersson et al., Bioseparation 2:15-22 (1991).
Andersson et al., Cancer Res. 47:3624-3626 (1987).
Abstract No. 88059400, Database Medline, (Aug. 21, 1987).
Abstract No. 92207443, Database Medline, (Sep.-Oct. 1991).

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