EP1165507A1 - Biocatalyseur et son utilisation dans la r solution enzymatique de b ta-lactame rac mique - Google Patents

Biocatalyseur et son utilisation dans la r solution enzymatique de b ta-lactame rac mique

Info

Publication number
EP1165507A1
EP1165507A1 EP00918984A EP00918984A EP1165507A1 EP 1165507 A1 EP1165507 A1 EP 1165507A1 EP 00918984 A EP00918984 A EP 00918984A EP 00918984 A EP00918984 A EP 00918984A EP 1165507 A1 EP1165507 A1 EP 1165507A1
Authority
EP
European Patent Office
Prior art keywords
lactam
process according
enantiomerically enriched
ncimb
accession
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP00918984A
Other languages
German (de)
English (en)
Inventor
Stephen John Clifford Chirotech Tech. Ltd. TAYLOR
Philip Alexander Chirotech Technology Ltd. KEENE
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Chirotech Technology Ltd
Original Assignee
Chirotech Technology Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Chirotech Technology Ltd filed Critical Chirotech Technology Ltd
Publication of EP1165507A1 publication Critical patent/EP1165507A1/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D205/00—Heterocyclic compounds containing four-membered rings with one nitrogen atom as the only ring hetero atom
    • C07D205/12—Heterocyclic compounds containing four-membered rings with one nitrogen atom as the only ring hetero atom condensed with carbocyclic rings or ring systems
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/78—Hydrolases (3) acting on carbon to nitrogen bonds other than peptide bonds (3.5)
    • C12N9/86—Hydrolases (3) acting on carbon to nitrogen bonds other than peptide bonds (3.5) acting on amide bonds in cyclic amides, e.g. penicillinase (3.5.2)

Definitions

  • This invention relates to a process for the production of optically active ⁇ -lactams by enzymatic resolution of the racemic ⁇ -lactam.
  • ⁇ -lactam ring as a structural feature in biologically important compounds is well established, as is the synthetic utility of ⁇ -lactams.
  • the utility of ⁇ -lactams as masked ⁇ -amino acids is illustrated by their use as precursors to the C-13 taxol side chain.
  • the active molecules are often chiral, and it is preferable that they are accessible in the form of single enantiomers. This feature is also utilised in the design of stereodefined scaffolds, to generate single enantiomer compounds libraries for initial lead identification as part of a drug discovery programme.
  • Alicyclic ⁇ -amino acids can be cyclised to the corresponding ⁇ -lactams, typically by use of carbod ⁇ mide reagents. They are also compounds having biological or pharmaceutical activity.
  • the natural product cispentacin, (-)-(lR,2S)-2- aminocyclopentanecarboxylic acid has potent antijfungal properties. It is synthesised as a natural product in Streptomyces setonii and Bacillus cereus, and is inhibitory to strains of Candida.
  • the same amino acid has also been used to probe the relationship between structure and taste in L-aspartyl dipeptides, where the absolute structure of the ⁇ -amino acid strongly affected the taste of the dipeptide.
  • Optically pure alicyclic ⁇ -amino acids may be made, for example, by the enzyme- catalysed solvent-based bioresolution of the amino carboxyl esters (Kanerva et al. Tetrahedron: Asymmetry, 1996, 7, 1705).
  • Such racemic amino esters are readily synthesised from ⁇ -lactams of formula (1), which are themselves synthesised by cycloaddition of the appropriate cyclic alkene and chlorosulfonyl isocyanate, followed by reductive work-up using sodium sulfite:
  • WO-A-97/10713 and EP-A-0232017 disclose various bicyclic compounds, including 7-azabicyclo[4.2.0]oct-4-en-8-one of the formula
  • WO-A-97/10713 includes reference to enantiomers, but there is no evident means of resolution. For example, the compounds will not readily form salts with chiral resolving agents.
  • EP-A-0002564 discloses ⁇ -lactams, including bicyciic structures. It is suggested that the racemic lactams might be resolved, and refers to GB-A- 1273278, but that depends on melt crystallisation, and probably requires that the substrate exists as a conglomerate.
  • the present invention is based on the discovery of a novel lactamase biocatalyst that allows efficient access to a range of synthetically useful ⁇ -lactams and the corresponding ⁇ -amino acids.
  • a preferred embodiment of the invention is the application of this methodology to the preparation of novel single enantiomer cyclohexene-fused ⁇ - lactams.
  • the novel biocatalyst is capable of enantioselective hydrolysis of racemic lactams of the formulae (2) and (3)
  • a further aspect of the invention lies in the novel, enantiomerically enriched compounds of formulae (2) and (3), preferably as the levorotatory enantiomer.
  • Lactams (2) and (3) are conveniently synthesised by cycloaddition of chlorosulfonyl isocyanate with norbornadiene and cyclohexa-l,3-diene respectively (Stajer et al, Tetrahedron, 1984, 40, 2385; Malpass et al, J. Chem. Soc, Perkin Trans. 1, 1991, 2276).
  • Enantiomerically enriched products obtained from the bioresolution of lactams (2) and (3) can be used to prepare information-rich chiral scaffolds for elaboration into single enantiomer compounds libraries.
  • Thealkene functionality is amenable to a range of transformations, especially through reaction with oxidants.
  • conversion of (3) to the chiral scaffold (4) gives three points for structural elaboration into defined regions of 3-dimensional space.
  • the more rigid scaffold (5) can be prepared via bioresolution of (2).
  • the present invention embodies a novel ⁇ -lactamase biocatalyst from Pseudomonas putida, combining the key attributes of high enantioselectivity with superior catalytic activity to ⁇ -lactamases described previously.
  • This biocatalyst has been deposited (see Example 1). Use of this biocatalyst allows certain single enantiomer ⁇ - lactams to be prepared for the first time in synthetically useful amounts.
  • the biocatalyst can be used under conditions that can readily be determined, to produce a mixture of compounds. These can be separated by known methods. More particularly, the enantiomerically enriched ⁇ -amino acid produced by hydrolysis can be isolated. The isolated ⁇ -amino acid may then be subjected to a condensation reaction, to reform the ⁇ -lactam ring.
  • the biocatalyst can be used in whole cell or paste form.
  • the enzyme can also be isolated, by techniques known to those skilled in the art.
  • Examples 1 , 2, 4, 6 and 8 illustrate the invention.
  • Examples 3 , 5 and 7 illustrate the preparation of racemic substrates.
  • TFAA is trifluoroacetic anhydride.
  • Glycerol stocks of 96 bacterial strains were used to inoculate 1.0ml Tryptone soya broth (Oxoid CM 129) per well in 2.2ml 96-well plates (Advanced Biotechnologies AB-0661). These were then shaken at 25°C on a Heidolph Titramax 1000 incubator at max rpm for 45 hours. The cells were harvested by centrifugation at 1 OOOg, 4°C, for 10 minutes, and the cell pastes were stored at -20°C.
  • a lO ⁇ l loopfiil of colony of CMC 103381 was used to inoculate seed flasks (100ml Tryptone soya broth (Oxoid CM 129) per 500ml Erlenmeyer flask). These were shaken at 25°C, 3 OOrpm in a temperature-controlled shaker (New Brunswick
  • the fermentation medium contained, per litre: 15g yeast extract (Oxoid L21), 8g KH 2 PO 4 , 7g K 2 HPO , lg MgSO 4 .7H 2 O, lg (NH 4 ) 2 SO 4 , 1ml trace elements solution, 1.0ml polypropylene glycol (Merck 29767 6Y), and 20g glucose.
  • the trace elements solution contained, per litre: 250ml cone.
  • the 15ml of buffered solution remaining from the bioresolution described above contain lg (7.1 mmol) of amino acid (maximum).
  • To the stirred buffer solution was slowly added, at IOC, 1.55g (7.1mmol) dibutyl dicarbonate dissolved in 20ml THF.
  • the pH of the solution was maintained at 9 by addition of 3M NaOH solution.
  • the reaction mixture was allowed to warm up to room temperature and left stirring overnight. The reaction was halted after 18 hours and THF was removed by rotary evaporator.
  • the aqueous reaction mixture was acidified to pH3 with 10% potassium hydrogen sulfate solution.
  • the mixture was then extracted with 3 x 50ml of ethyl acetate and the combined organic extracts were dried over magnesium sulfate. Removal of solvent by rotary evaporator yielded 1.1 g, 62% of a white solid.
  • Cyclopentadiene was freshly prepared by thermolysis of cyclopentadiene dimer.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Zoology (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Wood Science & Technology (AREA)
  • Microbiology (AREA)
  • Biotechnology (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

Procédé de préparation d'un β-lactame enrichi énantiomèriquement, caractérisé en ce qu'il comprend l'hydrolyse énantiosélective du β-lactame correspondant en présence d'une lactamase capable d'effectuer l'hydrolyse énantiosélective de 3-azatricyclo[4.2.1.02,5]non-7-èn-4-one et de 7-azabicyclo[4.2.0]oct-4-èn-8-one.
EP00918984A 1999-03-31 2000-03-29 Biocatalyseur et son utilisation dans la r solution enzymatique de b ta-lactame rac mique Withdrawn EP1165507A1 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
GBGB9907415.5A GB9907415D0 (en) 1999-03-31 1999-03-31 Biocatalyst and its use
GB9907415 1999-03-31
PCT/GB2000/001198 WO2000058283A1 (fr) 1999-03-31 2000-03-29 Biocatalyseur et son utilisation dans la résolution enzymatique de béta-lactame racémique

Publications (1)

Publication Number Publication Date
EP1165507A1 true EP1165507A1 (fr) 2002-01-02

Family

ID=10850715

Family Applications (1)

Application Number Title Priority Date Filing Date
EP00918984A Withdrawn EP1165507A1 (fr) 1999-03-31 2000-03-29 Biocatalyseur et son utilisation dans la r solution enzymatique de b ta-lactame rac mique

Country Status (6)

Country Link
EP (1) EP1165507A1 (fr)
JP (1) JP2002539813A (fr)
AU (1) AU3974900A (fr)
CA (1) CA2360944A1 (fr)
GB (1) GB9907415D0 (fr)
WO (1) WO2000058283A1 (fr)

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE1807495A1 (de) * 1968-11-07 1970-06-11 Hoechst Ag Verfahren zur Trennung binaerer Gemische
DE2862419D1 (en) * 1977-11-12 1984-07-26 Beecham Group Plc Derivatives of 7-oxo-1-azabicyclo(3.2.0)-hept-2-ene-2-carboxylic acid, their preparation, pharmaceutical compositions containing them and intermediates
AU6849687A (en) * 1986-01-23 1987-08-14 Upjohn Company, The Antimicrobial n-acyl-2-azetidinones
ATE118208T1 (de) * 1989-10-16 1995-02-15 Chiroscience Ltd Chirale azabicycloheptanone und verfahren zu ihrer herstellung.
GB9108384D0 (en) * 1991-04-19 1991-06-05 Enzymatix Ltd Bicycloheptanes
AU6988996A (en) * 1995-09-21 1997-04-09 Novartis Ag 7-azabicyclo(4.2.0)oct-4-en-8-one derivates as plant-microbicides
DE69914458T2 (de) * 1998-02-17 2004-10-28 G.D. Searle & Co., Chicago Verfahren zur enzymatischen auflösung von laktamen

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO0058283A1 *

Also Published As

Publication number Publication date
WO2000058283A1 (fr) 2000-10-05
JP2002539813A (ja) 2002-11-26
AU3974900A (en) 2000-10-16
CA2360944A1 (fr) 2000-10-05
GB9907415D0 (en) 1999-05-26

Similar Documents

Publication Publication Date Title
Mollica et al. Synthesis of (S)-5, 6-dibromo-tryptophan derivatives as building blocks for peptide chemistry
Garcı́a-Urdiales et al. Enzymatic ammonolysis of ethyl (±)-4-chloro-3-hydroxybutanoate. Chemoenzymatic syntheses of both enantiomers of pyrrolidin-3-ol and 5-(chloromethyl)-1, 3-oxazolidin-2-one
US5284769A (en) Process for preparing a single enantiomer of a lactam using lactamase
JP6145538B2 (ja) 触媒を用いたシクロヘキサン−１，２−ジカルボン酸ジメチルの生物学的分割による（３ａＳ，７ａＲ）−ヘキサヒドロイソベンゾフラン−１（３Ｈ）−オンの製造方法
CA2161849A1 (fr) Production de composes optiquement actifs
KR100479894B1 (ko) 광학이성질적으로 보강된 ｎ-변형 락탐의 제조 방법
EP1165507A1 (fr) Biocatalyseur et son utilisation dans la r solution enzymatique de b ta-lactame rac mique
JP2010532992A (ja) ３，４−エポキシ酪酸エチルの微生物速度論的分割
EP1002871A1 (fr) Procédé pour la préparation de dérivés optiquement actifs de 3-hydroxy-pyrrolidine par hydroxylation enzymatique
JP4843813B2 (ja) 酵素を用いるＲ−体又はＳ−体のα−置換ヘテロサイクリックカルボン酸及びこれと反対鏡像の鏡像異性体のα−置換ヘテロサイクリックカルボン酸エステルの調製方法
Bertoli et al. Chemoenzymatic synthesis of optically active α-methylene-γ-carboxy-γ-lactams and γ-lactones
JP3650648B2 (ja) エナンチオマー純粋なラクタムの製法
JP2003534807A (ja) 酵素を使用するラセミα−置換ヘテロ環式カルボン酸の光学分割方法
Sigmund et al. Enantioselective enzymatic aminolysis of a racemic 2-isoxazolylacetate alkyl ester
EP0407190A2 (fr) Procédé de préparation d'intermédiaires antibactériens par hydrolyse enzymatique de substrats racémiques
AU2014200205B2 (en) Preparation of (3aS,7aR)-hexahydroisobenzofuran-1(3H)-one by catalyzed biological resolution of dimethyl cyclohexane-1,2-dicarboxylate
Tarui et al. Microbial enantioselective ester hydrolysis for the preparation of optically active 4, 1-benzoxazepine-3-acetic acid derivatives as squalene synthase inhibitors
Kamal et al. Lipase-catalyzed resolution of 1-chloro-3-[(4-morpholin-4-yl-1, 2, 5-thiadiazole-3-yl) oxy] propan-2-ol: Synthesis of (R)-and (S)-timolol
Klempier et al. Nitrilase‐and Nitrile Hydratase‐Catalyzed Enantioselective Preparation of Non‐Proteinogenic Amino Acids
MXPA99011966A (en) Process for preparing enantiomerically enriched n-derivatised lactams

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 20011015

AK Designated contracting states

Kind code of ref document: A1

Designated state(s): AT BE CH CY DE DK ES FI FR GB GR IE IT LI LU MC NL PT SE

AX Request for extension of the european patent

Free format text: AL;LT;LV;MK;RO;SI

17Q First examination report despatched

Effective date: 20021025

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN

18D Application deemed to be withdrawn

Effective date: 20040527