US4307192A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Cell-free synthesis of deacetoxycephalosporin C
Assignee: Massachusetts Institute of Technology
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Filing: 1981-01-07
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Granted: 1981-12-22
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Expires: 2001-01-07
Abstract
Abstract A cell-free process for converting isopenicillin N, delta -(L- alpha -aminoadipyl)-L-cysteinyl-D-valine (hereinafter "LLD") and 6-substituted derivatives thereof to deacetoxycephalosporin C (DACPC) by the following reaction sequence: <IMAGE> is disclosed. In addition to the starting material, the reaction system includes ATP and a fresh extract of Cephalosporium acremonium prepared and used in a manner to preserve the racemase agent or agents necessary for conversion of the isopenicillin N to penicillin N, a necessary intermediate step in the process.
Inventors
- 👤 Arnold L. Demain
- 👤 Toshio Konomi
- 👤 Jack E. Baldwin
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-07 05:27
CPC Classifications
C; C12; C12P; C12P35/00; C12P35/06; C12P37/00; Y; Y10; Y10S; Y10S435/00; Y10S435/8215; Y10S435/911; Y10S435/925; Y10S435/926
Claims Analysis
Total Claims: 10
Claim 1
Independent Claim
158 words
A process for producing deacetoxycephalosporin C or 6-substituted derivatives thereof comprising the steps of: A. providing a starting material selected from the group consisting of: (a) a derivative of L-α-aminoadipyl-L cysteinyl-D-valine of the formula: ##STR8## and (b) an isopenicillin derivative of the formula: ##STR9## where R is selected from the group consisting of hydrogen, methyl, ethyl, propyl, and isopropyl; B. preparing a cell-free extract of Cephalosporium acremonium, the preparation of the extract being such as to preserve the operability of an enzyme contained therein capable of inverting the aminoadipyl side chain of the penicillin molecule from the L to the D conformation; C. contacting the extract and the starting material in a reaction zone while promoting oxygen transfer; D. providing ATP as an energy source to said reaction zone; and E. allowing a component of the extract to react with said starting material for a sufficient amount of time to produce a cephalosporin compound of the formula: ##STR10##
Claim 2
Dependent Claim
158 words
A process for producing deacetoxycephalosporin C or 6-substituted derivatives thereof comprising the steps of: A. providing a starting material selected from the group consisting of: (a) a derivative of L-α-aminoadipyl-L cysteinyl-D-valine of the formula: ##STR8## and (b) an isopenicillin derivative of the formula: ##STR9## where R is selected from the group consisting of hydrogen, methyl, ethyl, propyl, and isopropyl; B. preparing a cell-free extract of Cephalosporium acremonium, the preparation of the extract being such as to preserve the operability of an enzyme contained therein capable of inverting the aminoadipyl side chain of the penicillin molecule from the L to the D conformation; C. contacting the extract and the starting material in a reaction zone while promoting oxygen transfer; D. providing ATP as an energy source to said reaction zone; and E. allowing a component of the extract to react with said starting material for a sufficient amount of time to produce a cephalosporin compound of the formula: ##STR10##
Claim 3
Dependent Claim
12 words
The process as set forth in claim 1 wherein R is hydrogen.
Claim 4
Dependent Claim
21 words
The process as set forth in claim 1 wherein the extract used in step C has not been homogenized or frozen.
Claim 5
Dependent Claim
28 words
The process as set forth in claim 1 wherein the ATP utilized is regenerated by a salt-free ATP regenerating system comprising a phosphate donor and a phosphotransferease enzyme.
Claim 6
Dependent Claim
22 words
The process as set forth in claim 4 wherein the phosphate donor is phosphoenol pyruvate and the phosphotransferase enzyme is pyruvate kinase.
Claim 7
Dependent Claim
27 words
The process as set forth in claim 1 wherein said cell-free extract is a fresh extract prepared by treating a Cephalosporium acremonium culture with a lysing enzyme.
Claim 8
Dependent Claim
28 words
The process as set forth in claim 6 wherein said cell-free extract is made by treating Cephalosporium acremonium cells with endo β(1→3) glucanase, endo β(1→4) glucanase, and zymolyase.
Claim 9
Dependent Claim
22 words
The process as set forth in claim 1 wherein oxygen transfer is promoted by shaking the reaction components in the reaction zone.
Claim 10
Dependent Claim
33 words
The process as set forth in claim 1 wherein sucrose and trace concentrations of KCl and MgSO 4 are included in the reaction system, and the system is buffered to about pH 7.2.
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) | 1984-12-21 | 1985-06-22 | 1985-12-22 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1988-12-21 | 1989-06-22 | 1989-12-22 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1992-12-21 | 1993-06-22 | 1993-12-22 | $7700.00 | Window Closed / Lapsed |