US4510246A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Isolation of cyclase, epimerase and a ring expansion enzyme for producing unnatural cephalosporins
Assignee: Queen's University
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Filing: 1982-08-23
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Granted: 1985-04-09
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Expires: 2002-08-23
Abstract
Abstract The enzymes, cyclase, epimerase and a ring expansion enzyme, are isolated separately from a cell-free extract of a prokaryotic beta-lactam producing organism. The isolated enzymes are used to produce unnatural cephalosporins from polypeptide precursors. Isolation is carried out by adding ammonium sulfate to 40% saturation to the cell-free extract to precipitate contaminating proteins, adding ammonium sulfate to 70% saturation to the resultant supernatant to precipitate the enzymes, suspending the precipitated enzymes in a buffer, separating epimerase from the suspension by gel filtration, and separating cyclase and the ring expansion enzyme from each other by ion exchange chromatography.
Inventors
- 👤 Saul Wolfe
- 👤 Donald Westlake
- 👤 Susan Jensen
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-07 12:21
CPC Classifications
C; C12; C12N; C12N9/00; C12N11/00; C12N11/18; C12N9/90; C12P; C12P35/00; C12P35/06; C12P37/00; Y; Y10; Y10S; Y10S435/00; Y10S435/814; Y10S435/815; Y10S435/816
Claims Analysis
Total Claims: 4
Claim 1
Independent Claim
93 words
A process for isolating cyclase, epimerase and a ring expansion enzyme from a cell-free extract of a prokaryotic beta-lactam producing organism comprising: (a) precipitating contaminating proteins from said cell-free extract by addition of ammonium sulfate to 40% saturation; (b) separating precipitated protein from a supernatant; (c) adding ammonium sulfate to 70% saturation to said supernatant thereby precipitating desired enzymes; (d) suspending said precipitated desired enzymes in pH 7 buffer; (e) separating the epimerase by gel filtration chromatography; and (f) separating the cyclase and ring expansion enzymes from each other by ion exchange chromatography.
Claim 2
Dependent Claim
93 words
A process for isolating cyclase, epimerase and a ring expansion enzyme from a cell-free extract of a prokaryotic beta-lactam producing organism comprising: (a) precipitating contaminating proteins from said cell-free extract by addition of ammonium sulfate to 40% saturation; (b) separating precipitated protein from a supernatant; (c) adding ammonium sulfate to 70% saturation to said supernatant thereby precipitating desired enzymes; (d) suspending said precipitated desired enzymes in pH 7 buffer; (e) separating the epimerase by gel filtration chromatography; and (f) separating the cyclase and ring expansion enzymes from each other by ion exchange chromatography.
Claim 3
Dependent Claim
24 words
A process as claimed in claim 1 wherein said prokaryotic organism is selected from the group comprising S. clavuligerus, S. cattleya and S. lipmanii.
Claim 4
Dependent Claim
14 words
A process as claimed in claim 2 wherein said prokaryotic organism is S. clavuligerus.
References Cited (Backward Citations)
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1988-04-08 | 1988-10-08 | 1989-04-09 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1992-04-08 | 1992-10-08 | 1993-04-09 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1996-04-08 | 1996-10-08 | 1997-04-09 | $7700.00 | Window Closed / Lapsed |