Cyclization of .DELTA.-(L-.ALPHA.-aminoadipyl)-L-cysteinyl-D-valine to penicillins by cell-free extracts of Streptomyces clavuligerus.

Jensen, Susan E.; Westlake, D. W. S.; Wolfe, Saul

doi:10.7164/antibiotics.35.483

Cited by 75 publications

(33 citation statements)

References 21 publications

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“…The biosynthetic versatility of this organism, in terms of the synthesis of p-lactam compounds, has prompted an investigation of the enzymatic steps involved in the natural production of these antibiotics (3)(4)(5). Concurrent studies on the penicillin and cephalosporin-producing enzymes of Cephalosporium acremonium have been undertaken by several groups, and continue at the present time (6)(7)(8).…”

Section: Introductionmentioning

confidence: 99%

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Penicillin formation by cell-free extracts of Streptomycesclavuligerus. Behaviour of aminoadipyl-modified analogs of the natural peptide precursor δ-(L-α-aminoadipyl)-L-cysteinyl-D-valine (ACV)

Jensen¹,

Westlake²,

Bowers³

et al. 1984

Can. J. Chem.

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. Can. J. Chem. 62, 27 12 (1984). Six analogs of ACV, the biosynthetic precursor of the penicillin nucleus, have been synthesized, in which the 6-(L-a-aminoadipyl) moiety has been replaced by : P-(L-aspartyl), y-(L-glutamyl), 6-(D-a-aminoadipyl), adipyl, glycyl-6-(L-a-aminoadipyl), and N-acetyl-6-(L-a-aminoadipyl). The penicillins having the adipyl, glycyl-6-(L-a-aminoadipyl), and N-acetyl-6-(L-a-aminoadipyl) side chains have also been synthesized. Several improvements have been made in earlier routes to ACV and related peptides, including a simplified preparation of a dipeptide precursor, and a new synthesis of a-aminoadipic acid from lysine. A new reagent, 2-acetoximino-2-phenylacetonitrile, has been synthesized, which allows ready N-acetylation of amino hydroxyacids in aqueous acetone at room temperature. The peptide analogs have been examined as substrates of the enzyme isopenicillin N synthetase, which converts ACV to isopenicillin N in the presence of oxygen, ferrous ions, and ascorbate. The enzyme has been isolated from the prokaryotic organism Streptomyces clavuligerus, and has been employed either as a crude salt precipitate or in semi-purified form, freed from other enzymes of the penicillin-cephalosporin pathway. With the crude enzyme preparation, three of the analogs are active substrates, viz., adipyl, glycyl-6-(L-a-aminoadipyl), and N-acetyl-6-(L-a-aminoadipyl), but the latter two are converted, only via ACV, to isopenicillin N, the normal cyclization product. That the crude enzyme preparation contains a protease that deacylates N-substituted ACV analogs to ACV is confinned, inter alia, by the behaviour of the purified enzyme; of the various analogs, only the adipyl compound is an active substrate, but the conversion of this analog to carboxybutylpenicillin proceeds with only 1 -2%.efficiency, compared to the natural substrate. On a synthCtisC six analogues de I'ACV, le precurseur biosynthttique du noyau de la pknicilline, dans lesquel on a remplacC I'unitC 6-(L-a-aminoadipyl) par les unitts suivantes: P-(L-aspartyl), y-(L-glutamyl), 6-(D-a-aminoadipyl), adipyl, glycyl-6-(L-a-aminoadipyl) et N-acCtyl-6-(L-a-aminoadipyl). On a Cgalement synthCtisC des ptnicillines ayant des groupes adipyl, glycyl-6-(L-a-aminoadipyl) et N-acCtyl-6-(L-a-aminoadipyl) comme chaines lattrales. On a apportt plusieurs ameliorations aux voies d'accts a 1'ACV et aux peptides apparent& que nous avions dtcrites anttrieurement; ces amiliorations comprennent entre autre une prkparation simplifiCe d'un dipeptide prkcurseur ainsi qu'une nouvelle synthtse de I'acide a-aminoadipique a partir de la lysine. On a synthCtisC un nouveau reactif, I'acCtoxymino-2 phenyl-2 acitonitrile, qui permet d'effectuer facilement une N-acCtylation des hydroxy-acides amints en opirant dans l'acttone aqueuse et a la temperature ambiante. On a CtudiC le comportement des analogues peptidiques comme substrats de l'enzyme isopCnicilline N-synthktase qui transforme I'ACV en iso@nicilline N en presence d'oxygkne, d'ions ferreux et d'ions ascorbate. On a isol...

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Section: Introductionmentioning

confidence: 99%

“…However, it has since become clear (3)(4)(5) that ACV is a precursor of both penicillins and cephalosporins. The biosynthetic significance of GACV therefore remains unknown.…”

Section: Introductionmentioning

confidence: 99%

Penicillin formation by cell-free extracts of Streptomycesclavuligerus. Behaviour of aminoadipyl-modified analogs of the natural peptide precursor δ-(L-α-aminoadipyl)-L-cysteinyl-D-valine (ACV)

Jensen¹,

Westlake²,

Bowers³

et al. 1984

Can. J. Chem.

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“…chrysogenum TD189, a disrupted strain lacking isopenicillin epimerase activity, 15 and P. chrysogenum strains ABC and npe6 pyrG, both lacking the penDE gene, 10,16,17 were used. Micrococcus luteus ATCC 9341, an IPN sensitive bacterial strain, [18][19][20] was used for bioassays in each step of purification. b-lactamase from Bacillus cereus UL1 was used to check the presence of penicillin compound in these assays.…”

Section: Methods Microorganismsmentioning

confidence: 99%

A preparative method for the purification of isopenicillin N from genetically blocked Acremonium chrysogenum strain TD189: studies on the degradation kinetics and storage conditions

et al. 2011

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A protocol for preparative isopenicillin N (IPN) purification, a highly interesting and hitherto unavailable intermediate of the penicillin and cephalosporin biosynthetic pathway due to its high unstability, is described. Culture broths of Acremonium chrysogenum TD189, a strain blocked in cephalosporin biosynthesis that accumulates this metabolite, were treated with acetone and filtered though charcoal and a hydrophobic resin in a single step as tandem columns. The cleared broth was then lyophilized and passed though a Sephadex G-25 column. The last step was the purification to homogeneity of IPN in a semipreparative HPLC equipment and, optionally, a desalting step by Sephadex G-10 column. Once purified, a complete analysis of the stability of the compound and the conditions for its long-term storage was carried out. Our results suggest a first-order model for IPN decomposition for all the pH and temperature analyzed. IPN is more stable at neutral pH, and once lyophilized, can be stored under vacuum and À75 1C with a half-life of 770 days.

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“…Streptomyces clavuligerus NRRL 3585 and other Streptomyces cultures were maintained on sporulation medium as described previously (Jensen et al, 1982). For mutagenesis, S .…”

Section: Methodsmentioning

confidence: 99%

Discovery of an insertion sequence, IS116, from Streptomyces clavuligerus and its relatedness to other transposable elements from actinomycetes

Leskiw

Mevarech

Barritt

et al. 1990

Journal of General Microbiology

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We have identified an insertion sequence, IS116, present in Streptomyces clavuligerus at one copy per genome. The element was discovered as a 1.4 kb insertion into the multicopy plasmid pIJ702 after propagation in S. clavuligerus. The nucleotide sequence of IS116 and the flanking sequences from pIJ702 have been determined. The junctions with pIJ702 show no target site duplication and there are no inverted repeats at the ends of the element. One putative coding open reading frame of 1197 bp was identified which would code for a protein product of 399 amino acids. This protein resembles deduced integrase/transposase proteins specified by three other transposable elements of actinomycetes: IS110 and the mini-circle from Stveptomyces coelicolor A3(2), and -most particularly -IS900 of Mycobacterium paratuberculosis. Two regions that are relatively conserved among these gene products show features found in similar positions in many reverse transcriptases. IS116 and IS900 are also closely similar in their general organization and (apparently) in their insertion site specificity, whereas IS110 and the mini-circle are quite different in these features.

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Cyclization of .DELTA.-(L-.ALPHA.-aminoadipyl)-L-cysteinyl-D-valine to penicillins by cell-free extracts of Streptomyces clavuligerus.

Cited by 75 publications

References 21 publications

Penicillin formation by cell-free extracts of Streptomycesclavuligerus. Behaviour of aminoadipyl-modified analogs of the natural peptide precursor δ-(L-α-aminoadipyl)-L-cysteinyl-D-valine (ACV)

Penicillin formation by cell-free extracts of Streptomycesclavuligerus. Behaviour of aminoadipyl-modified analogs of the natural peptide precursor δ-(L-α-aminoadipyl)-L-cysteinyl-D-valine (ACV)

A preparative method for the purification of isopenicillin N from genetically blocked Acremonium chrysogenum strain TD189: studies on the degradation kinetics and storage conditions

Discovery of an insertion sequence, IS116, from Streptomyces clavuligerus and its relatedness to other transposable elements from actinomycetes

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