Abstract: The present invention is a novel method for maintaining and selecting recombinant DNA-containing host cells wherein the DNA encoding a selectable phenotype and the DNA encoding a useful polypeptide are the same. The aforementioned DNA is useful for expressing .delta.-aminolevulinic acid synthetase (ALAS) for the ultimate expression of .delta.-aminolevulinic acid (ALA) in yeast and related organisms. The invention further comprises plasmids pIT300, pIT301, pIT302, pIT304, pIT305, pIT306 and related Saccharomyces ALA deficient transformants. ALA is a five carbon amino acid that is useful as a light dependent herbicide.

Abstract: Site-specific mutagenesis of a gene for angiogenin producing DNA sequences encoding mutant proteins having increased angiogenic activity are disclosed. Expression vectors containing these sequences are introduced into host cells and direct the production of the mutant angiogenic proteins with markedly increased angiogenic and ribonucleolytic activity. Replacement of a single amino acid, the aspartic acid at position 116 of human angiogenin, with another amino acid including asparagine, alanine or histidine, yields mutant proteins with 8 to 15 fold increased ribonucleolytic activity toward tRNA and rRNA and 10 to 100 fold increased angiogenic potency in the chorioallantoic membrane assay. The mutant angiogenin proteins of this invention are useful therapeutic compositions to promote the development of a hemovascular network in a mammal or to promote wound healing, in particular, healing of torn or traumatized fibrocartilage material.

Abstract: Practical amounts of tissue plasminogen activator (tPA) whether secreted by cells naturally producing it or prepared by recombinant means can be solubilized by providing, in aqueous medium at pH 5-14 8 a solubilizing amount of a basic amino acid optionally and preferably, in the presence of a citric acid moiety. The ability to solubilize tPA at concentrations of up to 50 mg is significant as an aid in permitting smaller volumes for purification and permitting single injections of the drug as opposed to intravenous administration.

Abstract: A method of protecting soluble proteins such that their biological activity is preserved after freezing by exposing the protein to an amino acid or trimethylamine-N-oxide and transition metal ion prior to freezing. The protected protein can then be thawed without denaturation or impairment of the protein's biological activity. The protein is preferably exposed to the amino acid or trimethylamine-N-oxide by placing it in a 25-100 mM aqueous solution of organic solute and 1 mM Zn.sup.+2. This method is especially effective in preserving the biological activity of fragile proteins such as the enzyme phosphofructokinase. The present method can be used to preserve pharmaceutically useful proteins in a frozen form for storage and distribution. The treated protein can be thawed and administered directly to a user without removing the cryoprotectant since the amino acid or oxide and trace amounts of many transition metal ions are nontoxic.

Abstract: Cloning and expression of DNA segments encoding bovine IL-.alpha., and processes for producing purified bovine IL-1.alpha. as a product of recombinant cell culture, are disclosed.

Abstract: Constructions and methods for mutagenesis of nucleic acids involve chemical mutagenesis of a cassette comprising a structural gene linked to a non-functional restorable fragment of a marker gene. Mutants are detected by screening for the presence of the reconstituted marker among the ligation products of the cassette to a vector containing the non-functional restorable remainder of the marker gene. Xylose isomerase mutants, characterized by a change from glu (GAG) to lys (AAG) at amino acid position 262 in the xylA protein of E. coli were obtained by partial marker cassette mutagenesis. These mutants enzymes exhibited twice the rate of isomerization of glucose to fructose exhibited by the wild type.

Abstract: A mammalian myeloma cell comprising a double-stranded DNA molecule in its genome containing a coding sequence encoding a non-immunoglobulin protein, a non-immunoglobulin promoter sequence adjacent to the 5' terminus of said coding sequence, and the 8-base pair nucleotide sequence 5'-ATTTGCAT-3' located 5' to the transcription initiation site of said promoter sequence. The DNA molecule may optionally contain an enhancer element. Methods of producing non-immunoglobulin protein and DNA molecules are also provided.

Abstract: The tlrC gene is a novel tylosin resistance-conferring gene isolated from Streptomyces fradiae and used to construct a number of cloning vectors for use in Streptomyces. One such cloning vector, plasmid pSKC10, can be obtained in S. fradiae JS87 under the accession number NRRL 18072. S. fradiae JS87 is the preferred host when the tlrC gene is used to select tylosin-resistant Streptomyces transformants.

Abstract: The invention relates to new microorganisms of the Phanerochaete chrysosporium strain, which are useful especially for the production of lignin-degrading enzyme.These new microorganisms are deposited under nos. C.N.C.M.-I-398 and C.N.C.M.-I-399.

Abstract: A purified thermostable enzyme is obtained that has unique characteristics. Preferably the enzyme is isolated from the Thermus aquaticus species and has a molecular weight of about 86,000-90,000 daltons. The thermostable enzyme may be native or recombinant and may be used in a temperature-cycling chain reaction wherein at least one nucleic acid sequence is amplified in quantity from an existing sequence with the aid of selected primers and nucleotide triphosphates. The enzyme is preferably stored in a buffer of non-ionic detergents that lends stability to the enzyme.

Abstract: A novel gene conferring resistance to spiramycin in Streptomyces and related organisms was cloned from a genomic library of Streptomyces ambofaciens DNA. A thirty-one Kb fragment of S. ambofaciens DNA including the spiramycin-resistance gene was isolated from this library on a cosmid designated pKC592. The novel spiramycin-resistance gene can be isolated on an .about.2.9 Kb BamHI fragment by subcloning restriction fragments obtained from the pKC592 insert DNA. This BamHI fragment contains all of the information required for the expression of the spiramycin resistant phenotype in Streptomyces. Vectors and transformants containing the novel spiramycin resistance gene are provided.

Abstract: A process for converting AMP into ATP which comprises (a) using an enzyme which converts AMP into ADP and has been produced from microorganisms having an optimum growth temperature of 50.degree. C. to 85.degree. C. and an enzyme which converts ADP into ATP and has been produced from microorganisms having an optimum growth temperature of 50.degree. to 85.degree. C. is disclosed. In addition, there is disclosed a process for producing a physiologically active substance by a multienzyme process which comprises forming ATP from AMP by the step (a), (b) synthesizing a physiologically active substance with the resulting ATP, coverting AMP resulting from the reaction in step (b) into ATP by the reaction in step (a), and repeatedly utilizing the converted ATP for synthesis of the physiologically active substance in step (b). By using the process it is possible to stably and efficiently carry out conversion of AMP into ATP over a long period of time.

Abstract: A chemically-synthesized oligonucleotide composing a portion of the nucleotide sequence of the human IL-2 is employed as a probe to isolate the gene coding for human IL-2. The human IL-2 gene is selected from a cDNA library prepared from RNA produced by mitogen-stimulated Jurkat cells. Double-stranded cDNA is prepared from polyadenylated RNA extracted from bovine cells thought to produce interleukin-2. Such cDNA is inserted within a plasmid vector and the recombinant plasmid employed to transform hosts. Plasmid DNA, prepared from pools of the transformed hosts, is hybridized with a probe composed of a large portion of the coding sequence of the human IL-2 gene. Pools of host cells that provide signal to the human cDNA probe are identified, subdivided, and rescreened until a single positive colony is identified. Bovine plasmid cDNA is prepared from this colony, and the bIL-2 gene is sequenced.

Abstract: Different from conventional uricase products, the uricase of the present invention has outstandingly high thermal stability and is active in a wide range of pH from 5 to 10 for the oxidative decomposition of uric acid undertaken in clinical analysis. The uricase of the invention is produced microbiologically by a thermophilic microorganism belonging to the genus of Bacillus and especially named as Bacillus sp. TB-90 which is a novel species distinguishable from any of the microorganisms belonging to the genus of Bacillus.

Abstract: The tlrB gene is a novel tylosin resistance-conferring gene isolated from Streptomyces fradiae and used to construct a number of cloning vectors for use in Streptomyces and related organisms. One such cloning vector, plasmid pSVB9, can be obtained in S. lividans under the accession number NRRL 18073. S. lividans is the preferred host when the tlrB gene is used to select tylosin-resistant Streptomyces transformants.

Abstract: A process for producing protease by cultivating a protease-producing mold in a liquid medium, which is characterized by continuously adding a liquid medium containing a protein material to the culture medium after the substantial termination of proliferation of the mold cells.

Abstract: A method is provided for effecting the reduction of PCB contamination in organic waste by utilizing a particular strain of a biologically pure culture of Alcaligenes eutrophus under aerobic conditions.

Abstract: Methods and compositions are provided for efficient expression of genes in unicellular microorganisms, particularly yeast. The systems involve an expression system employing transcriptional initiation regions from glycolytic enzymes, particularly a chimeric expression system, having a first region providing for regulatable or constitutive expression, a second region providing for transcriptional initiation, where regions one and two are not found joined together in functional relationship in nature, and optionally a sequence providing for a secretory leader and processing signal, where the expression cassette will be joined to a gene which may be homologous or heterologous to the host. The expression cassette can be used on an extrachromosomal element or integrated into the host genome, whereby continuous expression can be achieved or inducible expression is obtained, by virtue of the presence or absence of an inducer.

Abstract: The present invention provides a transcriptional and translational activating sequence derived from the lambda pL transcriptional activating sequence and the E. coli lpp translational activating sequence. The activating sequence has been cloned into recombinant DNA expression vectors into which DNA sequences encoding funtional polypeptides can be readily inserted and expressed. The activating sequence of the present invention has been shown to drive high-level expression of a bovine growth hormone derivative and a human growth hormone derivative in E. coli. Preferred expression vectors of the present invention also comprise the cI857 temperature-sensitive lambda pL repressor gene, a rop.sup.- derivative of the plasmid pBR322 replicon, and a tetracycline resistance-conferring gene.

Abstract: Disclosed is a heterodimeric T lymphocytes receptor subunit. The subunit consists of variable, joining, constant, transmembrane, and cytoplasmic regions.The structure, amino acid, and nucleotide sequence of the lymphocyte receptor subunit were determined using cDNA clones derived from a functional murine cytotoxic T lymphocyte clone. The genes corresponding to these cDNA are expressed and rearranged specifically in T cells and have significant sequence homologies to immunoglobulin V and C genes.T cell receptor subunits may be produced from the cDNA clones. The protein molecules may be further used for the production of T-cell clone specific antibodies.