EP0244147A2
Expired / Lapsed
EP
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Purification process for hybrid proteins
Assignee: Merck and Co Inc
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Filing: 1987-04-22
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Granted: —
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Expires: 2007-04-22
Abstract
Abstract A hybrid protein containing a β-galactosidase moiety linked to a desired protein through a renin cleavage site is adsorbed to an affinity matrix under conditions in which no or minimal nonspecific adsorption occurs. The matrix-hybrid protein complex is then washed with buffer to remove loosely bound or associated proteins and resuspended in incubation buffer and renin is added to cleave the hybrid protein from the complex. The desired protein is released to the supernatant liquid while the β-galactosidase moiety remains bound to the affinity matrix. Purification of the desired protein is accomplished by anion exchange chromatography.
Inventors
- 👤 Amanda M. Sha
- 👤 E. Dale Lehman
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-29 10:58
CPC Classifications
C; C12; C12P; C12P21/00; C12P21/02; C07; C07K; C07K14/00; C07K14/005; C12N; C12N15/00; C12N15/09; C12N15/11; C12N15/62; C07K2319/00; C07K2319/40; C07K2319/50; C07K2319/61; C12N2710/00; C12N2710/00011; C12N2710/16011; C12N2710/16211; C12N2710/16222
Claims Analysis
Total Claims: 15
Claim 1
Independent Claim
53 words
A method of purifying hybrid proteins containing a β-galactosidase moiety linked to a desired protein through a renin cleavage site comprising: a. attachment of the hybrid protein by the β-galactosidase moiety to an affinity matrix; and b. release of the desired protein from the hybrid protein by renin cleavage of the cleavage site.
Claim 2
Independent Claim
53 words
A method of purifying hybrid proteins containing a β-galactosidase moiety linked to a desired protein through a renin cleavage site comprising: a. attachment of the hybrid protein by the β-galactosidase moiety to an affinity matrix; and b. release of the desired protein from the hybrid protein by renin cleavage of the cleavage site.
Claim 3
Dependent Claim
21 words
A method according to Claim 1, step a, wherein the affinity matrix is p-amino-phenyl-β-D-thio-galactoside attached to agarose by a linking structure.
Claim 4
Dependent Claim
14 words
A method according to Claim 2 wherein the linking structure contains 7 carbon atoms.
Claim 5
Dependent Claim
14 words
A method according to Claim 2 wherein the linking structure has a positive charge.
Claim 6
Dependent Claim
24 words
A method according to Claim 2 wherein the linking structure is a compound of formula: in which A represents agarose and T represents p-amino-phenyl-β-D-thio-galactoside.
Claim 7
Dependent Claim
14 words
A method according to Claim 2 wherein the agarose is cross linked agarose gel.
Claim 8
Dependent Claim
23 words
A method according to Claim 1 wherein the desired protein is separated from the renin by anion exchange chromatography and an acceptable eluant.
Claim 9
Dependent Claim
13 words
A method according to Claim 7, where the anion exchanger is trimethylammonium methyl.
Claim 10
Dependent Claim
16 words
A method according to Claim 7 wherein the desired protein is eluted with a salt gradient.
Claim 11
Dependent Claim
12 words
A method according to Claim 9 wherein the salt is sodium chloride,.
Claim 12
Dependent Claim
15 words
A method according to Claim 7 wherein the desired peptide is Epstein Barr Virus-Membrane antigen.
Claim 13
Dependent Claim
34 words
A composition comprising a desired peptide attached to β-galactosidase by a renin cleavage site bound to a ligand, p-aminophenyl-β-D-thio-galactoside which in turn is attached to agarose by a 7 carbon positively charged linking structrure.
Claim 14
Dependent Claim
34 words
A composition comprising a desired peptide attached to β-galactosidase by a renin cleavage site bound to a ligand, p-aminophenyl-β-D-thio-galactoside which in turn is attached to agarose by a 7 carbon positively charged linking structrure.
Claim 15
Dependent Claim
37 words
A composition comprising the desired peptide of Claim 11 attached to β-galactosidase by a renin cleavage site bound to the ligand, p-amino-phenyl-β-D-thio-galactoside which in turn is attached to agarose by a 7 carbon positively charged linking structure.
References Cited (Backward Citations)
No citation records indexed.
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