Eli Lilly and Company

Stephen W Queener

Paul Luther Skatrud

Thomas D Ingolia

Joseph M Zock

Cathleen Alice Cantwell

Roland L Hodges

Suellen M Samson

Otis W Godfrey

JoAnn Hoskins

Claude H Nash, Iii

Kevin R Kaster

Lucinda G Carr

Derek McGilvray

James R Swartling

Jerry L Chapman, Jr

Barbara J Wiegel

The present invention provides DNA compounds that encode ACV synthetase activity of,. The compounds can be used to construct recombinant DNA expression vectors for a wide variety of host cells, including,, Penicillium, Cephalosporium, and Aspergillus.

Recombinant DNA compounds that encode ACV synthetase activity of Cephalosporium acremonium

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

Recombinant DNA expression vectors and DNA compounds that encode deacetoxycephalosporin C synthetase and deacetylcephalosporin C synthetase

Heterologous extra-cellular expression of recombinant proteins in soluble functional form is desirable because of the ease associated with purification of the secreted proteins and avoidance of the need for cell extraction and protein refolding procedures. The present invention provides DNA sequences of the naturally-occurring phthalyl amidase gene isolated from Xanthobacter agilis that control transcription, translation, and extra-cellular secretion of proteins in Streptomyces lividans. These DNA sequences can be used in a method for extra-cellular expression of a wide variety of proteins in soluble functional form.

Method for extra-cellular expression of protein

Phthalyl amidase is an enzyme previously unknown in the art that catalyzes removal of the phthalyl moiety from phthalyl-containing amides. The current invention provides DNA compounds encoding the phthalyl amidase enzyme and methods for expressing such compounds. The present invention also provides recombinant DNA vectors encoding phthalyl amidase and host cells transformed with these DNA vectors.

Genes encoding and method of expressing a novel enzyme: phthalyl amidase

The present invention provides isolated DNA compounds and recombinant DNA cloning and expression vectors that encode PNB esterase from Bacillus subtilis. The invention also provides host cells transformed with these vectors and a method for production of the PNB esterase by recombinant DNA techniques.

Recombinant DNA compounds and expression vectors encoding

Genes encoding and method of expressing an enzyme: phthalyl amidase

The present invention comprises novel DNA compounds that encode isopenicillin N synthetase. The invention also comprises methods, transformants, and polypeptides related to the novel DNA compounds. The novel isopenicillin N synthetase-encoding DNA, together with its associated transcription and translation activating sequence, was isolated from Penicillium chrysogenum. The isopenicillin N synthetase-encoding DNA can be used to construct novel E. coli expression vectors that drive expression of isopenicillin N synthetase in E. coli. The intact P. chrysogenum isopenicillin N synthetase-encoding DNA and associated transcription and translation activating sequence can also be used to construct expression vectors that drive expression of the isopenicillin N synthetase in P. chrysogenum and Cephalosporium acremonium. The transcription and translation activating sequence can be fused to a hygromycin phosphotransferase-encoding DNA segment and placed onto expression vectors that function in P. chrysogenum and C. acremonium. The transcription termination and mRNA polyadenylation signals of the P. chrysogenum isopenicillin N synthetase can be used to increase ultimate expression of a product encoded on a recombinant DNA vector.

Recombinant DNA expression vectors and DNA compounds that encode

DNA compounds and recombinant DNA expression vectors that encode and drive expression in recombinant host cells of the isopenicillin N synthetase activity of Aspergillus nidulans are useful to produce isopenicillin N synthetase and to improve the yield of .beta.-lactam-containing antibiotics from antibiotic-producing organisms. The isopenicillin N synthetase gene of A. nidulans can be isolated from plasmid pOGOO4, available from the Northern Regional Research Center under the accession number NRRL B-18171.

The present invention comprises novel DNA compounds that encode isopenicillin N synthetase and also comprises related methods, transformants, and polypeptides. The novel isopenicillin N synthetase-encoding DNA, together with its associated transcriptional and translational activating sequence, was isolated from Cephalosporium acremonium and cloned into an E. coli cloning vector. The isopenicillin N synthetase-encoding DNA has been used to construct novel E. coli expression vectors that drive expression of a stable, active, and novel isopenicillin N synthetase in E. coli. The intact C. acremonium isopenicillin N synthetase-encoding DNA and associated transcriptional and translational activating sequence have also been used to construct C. acremonium expression vectors that drive expression of the isopenicillin N synthetase in C. acremonium. The C. acremonium transcriptional and translational activating sequence has further been fused to a hygromycin phosphotransferase-encoding DNA segment and placed onto C. acremonium expression vectors. Useful derivatives of the novel compounds and vectors are also described.

Recombinant DNA expression vectors and DNA compounds which encode

A method for transforming Cephalosporium and other lower eukaryotes is disclosed. The method involves inserting a recombinant DNA cloning vector comprising a Saccharomyces cerevisiae transcriptional and translational activating sequence positioned for expression of hygromycin phosphotransferase into a host cell and then growing the host cell under selective conditions. The vectors optionally further comprise Cephalosporium ribosomal DNA and also sequences that allow for replication and selection in E. coli and Streptomyces.

Vectors and conditions which allow genetic transformation of

A procedure for efficiently obtaining higher mycophenolic-acid-producing mutants from mycophenolic-acid-producing strains of the genus Penicillium by selecting mutants which are resistant to polyene antibiotics.

Growing community of inventors

Indianapolis, IN, United States of America

Stephen W Queener