Eniricerche S.p.a.

Snamprogetti

Enichem Synthesis S.p.a

Giuliano Galli

Guido Grandi

Renata Grifantini

Giuseppe Lucchese

Giovanna Carpani

Francesca De Ferra

Paola Cosmina

Gianni Frascotti

Paolo Carrera

Bruno Rappuoli

Carlo Di Gennaro

Barbara Camerini

Ada V Bellini

Ada Velati Bellini

Francesco Cecere

Angelo Tognoni

Gino Della Penna

A process is described for the production of D-&agr;-amino acids by the stereospecific conversion of racemic mixtures of 5-substituted hydantoins using a microorganism transformed with a plasmid which expresses an enzymatic system which converts the hydantoins into the corresponding D-&agr;-amino acids without the requirement for inducers. Also described is a plasmid containing genes encoding the enzymatic system, and a microorganism transformed with the plasmid.

Process for the production of D-&agr;-amino acids

The present invention relates to thermostable mutants of D-N-.alpha.-carbamoylase and means and methods for their preparation. The thermostable mutants have an unaltered or improved activity with respect to the wild-type enzyme and are particularly useful in the preparation of D-.alpha.-amino acids.

Thermostable Mutants of D-N-.alpha.-carbamoylase

The present invention relates to thermostable mutants of Agrobacterium radiobacter D-N-.alpha.-carbamoylase and means and methods for their preparation. The thermostable mutants have an unaltered or improved activity with respect to the wild-type enzyme and are particularly useful in the preparation of D-.alpha.-amino acids.

Thermostable mutants of D-N-.alpha.-carbamoylase

Mature Human Growth Hormone is prepared by enzymatic hydrolysis of a soluble precursor with immobilized Factor Xa. The soluble precursor contains a cleavage recognition sequence, Ile--Glu--Gly--Arg, for Factor Xa. The Factor Xa is preferably immobilized on CNBr activated Sepharose Cl-4B containing cyclic imido carbonate groups. Immobilization is carried out by reaction of amine groups on Factor X with the cyclic imido carbonate groups of the CNBr activated Sepharose Cl-4B to form covalently immobilized Factor X, which is then activated to Factor Xa with a protein contained in Russell's viper venom (RVV) in the presence of Ca.sup.++ ions.

Process for the preparation of mature human growth hormone by enzymatic

The present invention relates to mutants of D-N-.alpha.-carbamylase wherein at least one of the cysteines in position 243, 250 and 279 of the amino acid sequence of the wild type enzyme is substituted with a different residue selected from natural amino acids; a recombinant plasmid comprising a nucleotidic sequence which encodes for at least one of the mutants of D-N-.alpha.-carbamylase, host microorganisms transformed with said plasmid and a process for the preparation of these mutants by the culture of said microorganisms. The mutants of D-N-.alpha.-carbamylase have a higher enzymatic stability than that of the wild type enzyme and are particularly useful in the preparation of D-.alpha.-amino acids providing an improvement in the production yields.

Stable mutants of D-N-.alpha.-carbamoylase and process for preparing

In an improved process for the preparation of D-.alpha.-amino acids by the stereoselective conversion of racemic mixtures of 5-substituted hydantoins with an enzymatic system produced by a microorganism, the improvement consists in the fact that a microorganism is used which is transformed with the plasmid pSM700 cultivated at a temperature of between 20.degree. C. and 28.degree. C. The use of this microorganism allows an improvement in the expression of the enzymatic system and an increase in the conversion rate of the racemic hydantoin to D-.alpha.-amino acid.

Process for the preparation of D-.alpha.-amino acids

The present invention relates to mutants of D-N-.alpha.-carbamylase wherein at least one of the cysteines in position 243, 250 and 279 of the amino acid sequence of the wild type enzyme is substituted with a different residue selected from natural amino acids; a recombinant plasmid comprising a nucleotidic sequence which encodes for at least one of the mutants of D-N-.alpha.-carbamylase, host microorganisms transformed with said plasmid and a process for the preparation of these mutants by the culture of said microorganisms. The mutants of D-N-.alpha.-carbamoylase have a higher enzymatic stability than that of the wild type enzyme and are particularly useful in the preparation of D-.alpha.-amino acids providing an improvement in the production yields.

Nucleic acids encoding stable mutants of D-N-.alpha.-carbamoylase

A new plasmid vector is described which can replicate in B. subtilis and which can express and secrete, in the culture medium, human beta interleukin-1 (beta IL-1) without amino acid sequences extraneous to the natural molecule, a strain of B. subtilis transformed by the vector and a method for the expression and secretion of mature human beta interleukin-1. The human beta interleukin-1 thus obtained shows a specific acitvity of 1.times.10.sup.9 U/mg of protein and is particularly suitable as a stimulant for the immune system, as an adjuvant in vaccines for the activation of the repair mechanism in cases of tissue damage, and for the treatment of auto-immune diseases in man.

Expression and secretion of mature human beta interleukin-1 in Bacillus

Disclosed is a novel method for the in vivo and in vitro synthesis of modified peptides, using either engineered microorganisms capable of expressing peptide synthetases with modified substrate specificity, or peptide synthetases extracted from said microorganisms.

Method for genetically manipulating peptide synthetases

The structural gene encoding isoamylase from Pseudomonas SMP 1 is obtained. Also described are vectors comprising the isoamylase gene and host microorganisms transformed by these vectors. The transformed host microorganisms are capable of expressing and/or secreting mature isoamylase having biological activity.

Cloning of the gene coding the isoamylase enzyme and its use in the

A new plasmid vector is described which can replicate in B.subtilis and which can express and secrete, in the culture medium, human beta interleukin-1 (beta IL-1) without amino acid sequences extraneous to the natural molecule, a strain of B.subtilis transformed by the vector and a method for the expression and secretion of mature human beta interleukin-1. The human beta interleukin-1 thus obtained shows a specific acitvity of 1.times.10.sup.9 U/mg of protein and is particularly suitable as a stimulant for the immune system, as an adjuvant in vaccines for the activation of the repair mechanism in cases of tissue damage, and for the treatment of auto-immune diseases in man.

An enzymatic hydrolysis method is disclosed for the preparation of D-carbamyl aminoacids and the corresponding D-aminoacids. The method is based on the use of certain substituted hydantoins as the starting materials and these are hydrolized, in a temperature range from 10.degree. C to 70.degree. C, with hydropyrimidine hydrolase from calf livers. The pH range is from 8 to 9. The method in question is cheaper and more reliable than those of the prior art.

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Monterotondo, Italy

Giuliano Galli