Abstract: A mutated expandase enzyme having higher activity on penicillin G is provided to produce phenylacetyl-7-aminodeacetoxycephalosporanic acid (7-ADCA), which mutated expandase enzyme has one or more amino acid substitutions selected from M73T, G79E, V275I, L277K, C281Y, G300V, N304K, I305L and I305M.

Abstract: DNA sequences obtained from S. clavuligerus ATCC 27064, recombinant vectors incorporating such sequences and hosts transformed with such vectors are disclosed.

Abstract: The present invention relates to a process for delaying the deactivation of glutaryl amidase during enzyme catalysis. The enzyme catalyst is, after maximum conversion of the substrate, separated off by filtration and can be reused in a subsequent reaction batch. In each reaction cycle, the enzyme catalyst loses activity. The inactivation of the enzyme catalyst can be delayed by bringing it into contact with a thiol during the reaction.

Abstract: An enzymatic process for preparing 3-thiolated 7-aminocephalosporanic acid derivatives comprises the steps of enzymatically converting a 3-thiolated cephalosporin C of the formula I: to form a 3-thiolated-glutaryl-7-ACA of the formula II and enzymatically converting a compound of formula II to form a 3-thiolated-7-ACA of the formula III wherein R is a heterocyclic group comprising at least one nitrogen atom and R1 and R2 are both hydrogen atoms or one of them is a hydrogen atom and the other is an acyl donor.

Abstract: DNA sequences obtained from S. clavuligerus ATCC 27064, recombinant vectors incorporating such sequences and hosts transformed with such vectors are disclosed.

Abstract: The invention pertains to isolated phosphopantetheinyl transferases, such as the E. coli acyl carrier protein synthase, which transfer a phosphopantetheinyl group onto a substrate. The enzyme can be purified from a natural source, produced recombinantly, or synthetically. Accordingly, the invention provides compositions and kits including phosphopantetheinyl transferases and host cells expressing phosphopantetheinyl transferases. The invention also provides nucleic acids encoding phosphopantetheinyl transferases and vectors comprising such nucleic acids. The invention further provides methods for phosphopantetheinylating a substrate in vitro or in vivo and methods for producing antibiotics in vitro or in vivo.

Abstract: Methods are provided for increasing and altering the timing of antibiotic production in Streptomyces species, particularly S. coelicolor and S. lividans, by functionally deleting the S. coelicolor scbA and scbR genes, respectively, or their homologues. Also provided are strains having such mutations, and methods of producing antibiotics using such strains. Also provided are methods for identifying strains in which functional deletion of the scbA and/or scbR genes or their homologues leads to the above effects.

Abstract: Enzymatic process for the preparation of cephalosporanic 7?-(4-carboxybutananude) acid by using the modified enzyme D-aminoacid oxidase of Trigonopsis variabilis produced in Escherichia coli. The process for the expression of the enzyme comprises: (I) isolating the DNA corresponding to gene which codes for the enzyme D-aminoacid oxidase; (II) removing the intron which is contained in said gene; (III) inserting the DNA fragment obtained into the plasmide which is capable of replication in Escherichia coli; (IV) fusing at the extremity 5′ of the structural region of the gene a synthetic assembler which contains a nucleotide sequence which codes for six histidines; (V) transforming a strain of Escherichia coli with the resulting recombinant plasmide; (VI) cultivating the transformed cells of Escherichia coli; and (VII) recovering the enzyme D-aminoacid oxidase of the former cultivation operation through affinity chromatography.

Abstract: A process for preparing cephalosporanic acid derivatives comprises the steps of enzymatically converting a 3-thiolated cephalosporin C compound of formula III: 1

Abstract: An enzymatic process for preparing 3-thiolated 7-aminocephalosporanic acid derivatives comprises the steps of enzymatically converting a 3-thiolated cephalosporin C of the formula I: 1

Abstract: Disclosed is a process for the synthesis of &bgr;-lactam antibacterials using soluble side chain esters in the presence of enzyme acylase. Also disclosed are novel esters useful as reactants in said process.

Abstract: The present invention discloses a process for the production of N-deacylated cephalosporin compounds via the fermentative production of their 7-acylated counterparts.

Abstract: Methods for identifying organisms capable of degrading fumonisin. Fumonisin can be incorporated into culture medium for selection of organisms resistant to fumonisin and/or capable of growing on fumonisin as a sole carbon source. Using this method, several organisms have been identified. These organisms can be used to isolate the enzymes and the genes responsible for conferring fumonisin-resistance. The gene can be cloned and inserted into a suitable expression vector so that the protein can be further characterized. Additionally, the DNA encoding for fumonisin degrading enzymes can be used to transform plant cells normally susceptible to Fusarium or other toxin-producing fungus infection. Plants can be regenerated from the transformed plant cells. In this way, a transgenic plant can be produced with the capability of degrading fumonisin, as well as with the capability of producing the degrading enzymes.

Abstract: The invention relates to a process for the preparation of an antibiotic, in particular cefalexin, ampicillin, amoxicillin, cefaclor, cefradin, cefadroxil, cefotaxim and the like, wherein a beta-lactam core is acylated, the antibiotic is recovered from the reaction mixture, the remaining mother liquor is subjected to a hydrolysis reaction in which the antibiotic present in the mother liquor is decomposed into its initial compounds, in particular the beta-lactam core, and the acylation agent is hydrolized. The beta-lactam core can then be recovered virtually quantitatively or recycled to the acylation reaction. This is because it has been found that the solubility of the beta-lactam core is unexpectedly high at relatively high concentrations of acylation agent and antibiotic.

Abstract: A process for decreasing esterase activity present in the presence of D-amino acid oxidase activity or in the presence of glutarylacylase activity in a mixture having esterase activity and D-amino acid oxidase activity and/or having esterase activity and glutarylacylase activity by treatment with phenylmethylsulphonyl fluoride; immobilized biocatalyts in spherical particle form obtainable by treating microorganism cells having enzyme activity with a primary or secondary amine containing polymer, an organic solvent which is able to form a two-phase system with water and a bifunctional agent; and the use of that process and biocatalysts in the production of 7-aminocephalo-sporanic acid, 7-amino-3-hydroxymethyl-3-cephem-4-carboxylic acid and cephalosporin antibiotics.

Abstract: A process for the preparation of Cefuroxime acid (I), which comprises the following steps: (1) Extraction of deacetyl 7-glutaryl ACA (II) aqueous solution at acid pH with organic solvents (for example according to the procedures disclosed in U.S. Pat. No. 5,801,241); (2) drying the resulting solution while preventing lactonization of the intermediate; (3) carbamoylation of the hydroxymethyl group at the 3-position by reaction with chlorosulfonyl isocyanate or similar products; (7) extraction of the carbamoyl derivative from step 3 with water at neutral pH; (8) enzymatic hydrolysis of the amide at the 7-position of the cephalosporanic ring with glutaryl acylase; (6) acylation of the amnino group by condensation with 2-furanyl(sin-methoxyimino)acetic acid chloride or mixed anhydride.

Abstract: The present invention disclosures a process for the production of N-deacylated cephalosporin compounds via the fermentative production of their 7-acylated counterparts.

Abstract: A process is provided for delaying the deactivation of glutaryl amidase during catalysis. The amidase is, after maximum conversion of a substrate, separated off by filtration and can be reused in a subsequent reaction batch. In each reaction cycle, the amidase loses activity. The inactivation of the amidase can be delayed by bringing it into contact with a thiol during the reaction. The thiol is preferably 2-mercaptoethanol or cysteine.

Abstract: The present invention provides a biological system for expanding the dethiazolidine ring of penicillins into the dehydrothiazine ring of cephalosporins or cephalosporin precursors. In particular, the invention defines reaction conditions under which expandase enzyme can convert penicillin substrates other than penicillin N into cephalosporins. The invention therefore provides a relatively inexpensive, uncomplicated, and environmentally friendly biological system for cephalosporin production from penicillins, which system can replace the synthetic chemical approaches currently utilized. In particular, the invention provides a system for producing DOAG and/or DAG, which can be enzymatically converted into 7-ADCA and 7-ADAC, which, in turn, can be enzymatically or chemically converted into valuable cephalosporins of commerce.

Abstract: The present invention concerns the direct production of desacetylcephalosporin C by culturing a strain of Acremonium chrysogenum containing recombinant nucleic acid encoding Rhodosporidium toruloides cephalosporin esterase.

Abstract: The present invention discloses a process for the production of N-deacylated cephalosporin compounds via the fermentative production of their 7-acylated counterparts.

Abstract: The present invention relates to mutant D-amino acid aminotransferase, including nucleic acids encoding mutant D-amino acid aminotransferase. The mutant D-amino acid aminotransferase of the present invention is obtained by the substitution of the glutamate residue at position 13 of wild type D-amino acid aminotransferase from Bacillus sphaericus with hydrophobic amino acids. The mutant D-amino acid aminotransferase can be used in the production of D-amino acid and the conversion of glutaryl-7-aminocephalosporanic acid from cephalosporin C. The present invention also includes replica-paper staining method for screening the cells expressing high DAT activity.

Abstract: The present invention relates to a novel process for the preparation of cephalosporins having been deacylated at the 7-amino group, by fermentation of a cephalosporin producing microorganism in the presence of a side chain precursor, extraction of the N-substituted cephalosporin compound as present in the fermentation broth or fluid to an organic solvent, back extraction of the N-substituted cephalosporin compound to water, treatment of the aqueous phase with a dicarboxylate acylase and isolation of the crystalline cephalosporin compound according to formula (1) from the conversion solution, characterized in that the fermentation broth or fluid is incubated at acidic conditions and an elevated temperature prior to extraction of the N-substituted cephalosporin compound to an organic solvent.

Abstract: The present invention relates to an enzyme process for the one-step conversion of cephalosporin C or a derivative thereof into 7-aminocephalosporanic acid or a corresponding derivative thereof. The one step conversion is effected using a cephalosphorin C amidohydrolase derived from Pseudomonas Vesicularis B965, or from any cephalosporin C amidolydrolase producing or potentially producing descendants thereof, or from expression of DNA derived from Pseudomonas Vesicularis B965 or any cephalosporin C amidohydrolase producing or potentially producing descendants thereof.

Abstract: DNA encoding the gene for the synthetase enzyme capable of generating &dgr; (L-a-aminoadipyl)-L-crysteinyl-D-valine (ACV) from its constituent amino acids was obtained from several penicillin and cephalosporin producing organisms, e.g. Penicillium chrysogenum, cephalosporium and a Flavobacterium species. The DNA was used to prepare recombinant vectors comprising the ACV synthetase gene and hosts transformed with such vectors. The ACV synthetase gene can form part of a gene cluster comprising other genes involved in -&bgr;-lactam biosynthesis and the production of penicillin by expression of the entire biosynthetic gene cluster for the synthesis of penicillin from primary amino acids is described. Suitable hosts in which expression can take place include heterologous hosts which are naturally non-producers of penicillin.

Abstract: The present invention provides a biological system for expanding the dethiazolidine ring of penicillins into the dehydrothiazine ring of cephalosporins or cephalosporin precursors. In particular, the invention defines reaction conditions under which expandase enzyme can convert penicillin substrates other than penicillin N into cephalosporins. The invention therefore provides a relatively inexpensive, uncomplicated, and environmentally friendly biological system for cephalosporin production from penicillins, which system can replace the synthetic chemical approaches currently utilized. In particular, the invention provides a system for producing DOAG and/or DAG, which can be enzymatically converted into 7-ADCA and 7-ADAC, which, in turn, can be enzymatically or chemically converted into valuable cephalosporins of commerce.

Abstract: Beta-lactam antibiotics are synthesized by reacting an amino-beta-lactam component with a corresponding amino-group-containing acylating side-chain component in the presence of penicillin amidase from E. coli covalently immobilized on support particles. The resulting beta-lactam antibiotic product is solubilized by adding an acid such as sulfuric acid to lower the pH to 1.0 at a temperature in the range of 0° C. to +5° C. The immobilized penicillin amidase is substantially inactivated by the acid. After separating the beta-lactam antibiotic product, the immobilized penicillin amidase is substantially reactivated for reuse in antibiotic synthesis by treatment with a buffer having about a neutral pH. Antibiotics that can be produced include ampicillin, amoxicillin, cephalexin, cefaclor and cefadroxil.

Abstract: The present invention provides DNA compounds that encode the expandase/hydroxylase enzyme of Cephalosporium acremonium. The compounds can be used to construct recombinant DNA expression vectors for a variety of host cells, including E. coli, Penicillium, and Cephalosporium.

Abstract: A process for increasing the amount of clavam produced by an organism having both a clavam pathway or a portion thereof and a cephalosporin pathway or a portion thereof by interfering with the conversion of L-lysine to L-&agr;-aminoadipic acid in the cephalosporin pathway. Plasmids containing a defective LAT (lysine amino transferase) gene and organisms containing such plasmids are also provided.

Abstract: Disclosed is a process for the synthesis of .beta.-lactam antibacterials using soluble side chain esters in the presence of enzyme acylase. Also disclosed are novel esters useful as reactants in said process.

Abstract: A procedure for producing a modified 7.beta.-(4-carboxybutanamide) cephalosporinase enzyme which can be purified in a single chromatographic step. The procedure for production of the enzyme involves: mutagenizing the gene which codes for the enzyme from Acinetobacter sp. ATCC 53891 by inserting a nucleotide sequence coding for six histidine residues; fusing the mutant gene with high-efficiency promoter DNA sequences; transforming Escherichia coli cells with the fusion gene construct; growing the transformed Escherichia coli cells; and recovering the enzyme by the use of supports which contain metal chelates. 7-Aminocephalosporanic acid is an important intermediate for the manufacture of a wide range of antibacterial agents of the cephalosporin family.

Abstract: A bioprocess for preparing adipoyl-7-ACA comprising the steps: (a) transforming cells of a strain of Penicillium chrysogenum which produces isopenicillin N with an expression vector containing DNA encoding an enzyme, having expandase activity capable of accepting adipoyl 6-APA as a substrate, an enzyme having hydroxylase activity capable of accepting adipoyl-7-ADCA as a substrate and an enzyme having acetyl transferase activity capable of accepting adipoyl 7-ADAC as a substrate; (b) culturing the transformed cells from step a) in a suitable culture medium containing an adipate feedstock, wherein said cells produce adipoyl 6-APA; and (c) culturing the transformed cells producing adipoyl 6-APA of step b) under conditions suitable for expression of said DNA encoding enzyme, thereby producing the end product adipoyl-7-ACA.

Abstract: An enzymatic process for hydrolyzing the 7-position acyl ester of certain cephalosporin compounds to the corresponding alcohol is disclosed.

Abstract: A process is taught for the preparation and recovery of 7-aminodesacetoxycephalosporanic acid (7-ADCA) via enzymatic ring expansion activity on penicillin G, using a Penicillium chrysogenum transformant strain expressing expandase.

Abstract: A process for making 7-aminodeacetyl-cephalosporanic acid (7-ADAC).

Abstract: The present invention relates to the biosynthesis of .beta.-lactam antibiotics. More specifically, the invention relates to processes of producing .beta.-lactam antibiotics in vivo and in vitro. Also contemplated is a novel enzyme capable of catalyzing certain steps involved in .beta.-lactam biosynthesis. Further, the invention relates to a DNA construct encoding the novel enzyme, a recombinant vector or transformation vehicle comprising the DNA construct, and finally a cell comprising the DNA construct or recombinant vector.

Abstract: Disclosed is a process for the synthesis of .beta.-lactam antibacterials using soluble side chain esters in the presence of enzyme acylase. Also disclosed are novel esters useful as reactants in said process.

Abstract: The present invention relates to an enzyme process for the one-step conversion of cephalosporin C or a derivative thereof into 7-aminocephalosporanic acid or a corresponding derivative thereof.

Abstract: The disclosed process is for the recovery of cephalexin from a mixture containing cephalexin and 7-aminodesacetoxy cephalosporanic acid (7-ADCA), wherein a mixture of cephalexin and 7-ADCA, with a pH higher than 7, which apart from any solid cephalexin being present is homogeneous at a pH between 7 and 8.5, is subjected to a pH modification until a pH lower than 7.8 is reached, and the solid substance is recovered. The disclosed process is particularly suited for application to a reaction mixture obtained after the enzymatic acylation reaction of 7-ADCA with a phenylglycine derivative as an acylation agent. Pure cephalexin can thus be recovered in a simple manner.

Abstract: A method to produce novel polyketide structures by designing and introducing specified changes in the DNA governing the synthesis of the polyketide is disclosed. The biosynthesis of specific polyketide analogs is accomplished by genetic manipulation of a polyketide-producing microorganism by isolating a polyketide biosynthetic gene-containing DNA sequence, identifying enzymatic activities associated within the DNA sequence, introducing one or more specified changes into the DNA sequence which codes for one of the enzymatic activities which results in an altered DNA sequence, introducing the altered DNA sequence into the polyketide-producing microorganism to replace the original sequence, growing a culture of the altered microorganism under conditions suitable for the formation of the specific polyketide analog, and isolating the specific polyketide analog from the culture.

Abstract: The invention relates to an improved process for preparing cephalosporins by reaction compounds such as 7-amino-cephalosporanic acid, 7-amino-3-deacetoxy-cephalosporanic acid or their derivatives with derivatives of .alpha.-amino acids in the presence of a properly immobilized penicillin acylase enzyme under the following conditions, independently, or in combination: (1) at a temperature ranging from 0.degree. C. to +20.degree. C.; or (2) at ambient pH; with a high molar ratio of .alpha.-amino acid to a cephalosporanic nucleus.

Abstract: An overall efficient process for the preparation and recovery of 7-aminodesacetoxycephalosporanic acid (7-ADCA) via 3-(carboxyethylthio)propionyl-7-ADCA, using a Penicillium chrysogenum transformant strain expressing expandase in conjunction with acyltransferase, is provided.

Abstract: An enzyme selected from penicillin-G amidase, glutaryl-7-ACA acylase and D-amino acid oxidase is immobilized by covalent bonding on an aminofunctional organosiloxane polymer carrier to provide an immobilized enzyme having a specific volume activity of at least 100 U/g wet carrier. Preferably, the carrier has an average diameter or 0.01 to 3 mm and is essentially spherical. Covalent bonding is accomplished by activating amino groups on the carrier with a dialdehyde and reacting the activated groups with reactive groups on the enzyme. An amount of enzyme is immobilized to provide a weight ratio of enzyme to carrier of 1 to 300 mg protein per g wet carrier.

Abstract: D-amino acid oxidase (DAO) originated from Fusarium solani M-0718, a DNA compound encoding the oxidase, expression vectors containing said DNA compound are provided. Esherichia coli transformants capable of producing recombinant DAO in high level are also provided.

Abstract: A method for providing a semisynthetic .beta.-lactam antibiotic by enzyme catalyzed acylation of the parent .beta.-lactam with an activated derivative of the side chain acid wherein a modulator, which consists of one or more compounds different from the reactants and the reaction product and which suppresses the hydrolysis of the activated derivative of the side chain acid and the desired product more than it suppresses the synthesis of the desired product, is added to the reaction mixture, at the beginning of the reaction process, in a concentration from about 0.2 to 100.times.10.sup.3 .mu.m.

Abstract: A chemical analyzer has turn tables respectively carrying reaction vessels, sample containers and reagent containers disposed thereon in circular rows. A sample pipetting mechanism and a reagent pipetting mechanism are provided to transfer predetermined amounts of sample and reagent solution from a sample container and from a reagent container into a reaction vessel. The thus transferred sample and reagent solution are mixed in the reaction vessel by a stirrer comprising a piezoelectric element which is disposed in non-contacting relationship to the sample and reagent solution in the reaction vessel and is electrically energized to generate a sound wave which causes a circulating flow of the reagent solution in the reaction vessel whereby the sample and the reagent solution are mixed in a non-invasive manner.

Abstract: An overall process for the preparation and recovery of 7-aminodesacetoxycephalosporanic acid (7-ADCA) via enzymatic ring expansion activity on penicillin G, using a Penicillium chrysogenum transformant strain expressing expandase.

Abstract: An overall efficient process for the preparation and recovery of 7-aminodesacetoxycephalosporanic acid (7-ADCA) via 2-(carboxyethylthio)acetyl- and 3-(carboxymethylthio)propionyl-7-ADCA, using a Penicillium chrysogenum transformant strain expressing expandase in conjunction with acyltransferase, is provided.

Abstract: An enzyme preparation that exhibits cephalosporin chloroperoxidase activity is isolatable from a microorganism species of the Rathayibacter genus. This enzyme preparation can convert cephalexin to cefaclor in a single step. A particular, unique microorganism that can provide the cephalosporin chloroperoxidase enzyme preparation is Rathayibacter biopuresis.

Abstract: The present invention provides a process for producing 7-aminocephem compounds or salts thereof.