Cetus Corporation

Shing Chang

Hing C Wong

Arie Ben-Bassat

Keith A Bauer

Sheng-Yung Chang

David H Gelfand

James H Meade

Vaughan P Wittman

Method of obtaining N-terminal methionine-free proteins are described. The methods employ a novel enzyme, E. coli methionine aminopeptidase either in vitro or in vivo. For in vivo application, plasmid-borne DNA encoding the peptidase is transformed into a bacterial host which produces the desired protein.

Bacterial methionine n-terminal peptidase

The invention discloses modified signal peptides derived from wild-type signal peptides of the type that are capable of forming membrane-bound lipoproteins and methods for making such modified signal peptides and DNA sequences encoding them. Modified signal peptides of the invention and DNA sequences encoding them are useful for increasing the secretion of heterologous gene products produced by transformed host organisms. The invention further discloses a method for producing recombinant DNA sequences in vivo.

Modified signal peptides

A method and a cloning vector are described for the controlled accumulation of cloned heterologous gene products in Bacillus subtilis. The cloning vector is capable of being replicated in B. subtilis and includes the heterologous gene located and oriented such as to be under the control of an operator, promoter, and ribosome binding site sequence. The gene codes for a protein which is under the control of a transport mechanism by which the protein is secreted by the B. subtilis. The gene product is recovered from the growth medium for the B. subtilis. The cloning vector is also capable of similar use in other bacteria such as E. coli.

Method and vector organism for controlled accumulation of cloned

The invention concerns a method for extending the half-life of mRNAs. The half-life extension is conferred upon the mRNA by a co-transcribed positive retroregulatory element which is ligated to the 3' end of the DNA sequence encoding the RNA. RNAs having an extended half-life conferred by a co-transcribed positive retroregulatory element are also claimed.

3'-expression enhancing fragments and method

Methods of obtaining N-terminal methionine-free proteins are described. The methods employ a novel enzyme, E. coli methionine aminopeptidase either in vitro or in vivo. For in vivo application, plasmid-borne DNA encoding the peptidase is transformed into a bacterial host which produces the desired protein.

Bacterial methionine N-terminal peptidase

Methods for obtaining N-terminal methionine-free proteins involve a novel E. coli methionine amino peptidase. The method is capable of in vitro or in vivo application. For in vivo application, a plasmid-borne DNA encoding the peptide is expressed in a bacterial host.

Methods and compositions are described which can be used to enhance the stability of genetically engineered plasmids in Gram-positive organisms. Also described are genetically engineered plasmids comprising a DNA fragment which enhances their stability and Gram-positive organisms transformed with such plasmids.

Partition proficiency in genetically engineered plasmids for

The invention discloses a method and cloning vectors useful for the production of cloned heterologous gene products in B. subtilis. Use of the method and vector allows the host to produce the heterologous gene product as a single unfused peptide having no extraneous amino acids attached.

Method and vector organism for controlled accumulation of heterologous

The invention concerns positive retroregulatory elements, which when ligated to selected DNA sequences coding for a gene product, enhance the expression of the gene product. Plasmids carrying the positive retroregulatory element ligated to selected DNA sequence and cells transformed by such plasmids are provided AND claimed. In addition, the invention relates to a method for enhancing expression of a gene product by ligating a positive retroregulatory element to a selected DNA sequence expressionable for a desired gene product.

3'Expression enhancing fragments and method

A method and a cloning vector are described for the controlled accumulation of cloned heterologous gene products in Bacillus subtilis. The cloning vector is capable of being replicated in B. subtilis and includes the heterologous gene located and oriented such as to be under the control of an operator, promoter, and ribosomal binding site sequence. The gene codes for a protein which is under the control of a transport mechanism by which the protein is secreted by the B. subtilis. The gene product is recovered from the growth medium for the B. subtilis. The cloning vector is also capable of similar use in other bacteria such as E. coli.

The invention discloses a method for producing modified signal peptides sequences derived from wild-type signal peptide sequences of the type that are capable of forming membrane-bound lipoproteins. Modified signal peptide sequences produced by the method of the invention are useful for increasing the secretion of heterologous gene products produced by transformed host organisms. The invention further discloses a method for producing recombinant DNA sequences in vivo.

The invention is a plasmid vector in which a first DNA sequence expressionable for a selected gene product is ligated to a positive retroregulatory element in a relationship thereto whereby expression of the selected gene product is enhanced. The first DNA sequence is furthermore operably linked to a second and third DNA sequence operably linked to one another which are respectively the P.sub.L promoter and a DNA sequence corresponding to an N gene ribosome binding site. Microorganisms transformed with the plasmid are also claimed.

Polypeptide expression using a portable temperature sensitive control

Chimeric plasmids capable of transforming bacteria and yeast are described. The plasmids carry the Cm (chloramphenicol resistance) gene and the Tc (tetracycline resistance) gene as selectable markers. The Cm gene allows the plasmids to be selected for in wild-type strains of the yeast Saccharomyces cerevisiae. The Tc gene allows heterologous genes cloned into the plasmids to be selected for in Escherichia coli bacteria.

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Hercules, CA, United States of America

Shing Chang