Abstract: Disclosed herein are modified strains for reducing degradation of recombinantly expressed products secreted from a host organism and methods of using the modified strains. In some embodiments, to attenuate a protease activity in Pichia pastoris, the genes encoding enzymes the degrade proteases are inactivated or mutated to reduce or eliminate activity. In preferred strains, the protease activity of proteases encoded by PAS_chr4_0584 (YPS1-1) and PAS_chr3_1157 (YPS1-2) (e.g., polypeptides comprising SEQ ID NO: 66 and 67) is attenuated.

Abstract: DGA1 catalyzes the final enzymatic step for converting acyl-CoA and 1,2-diacylglycerol to triacylglycerols (TAG) and CoA in yeast. Disclosed are methods for expression in an oleaginous yeast host of polynucleotide sequences encoding DGA1 from Rhodosporidium toruloides, Lipomyces starkeyi, Aurantiochytrium limacinium, Aspergillus terreus, or Claviceps purpurea. Also described herein are engineered recombinant host cells of Yarrowia lipolytica comprising heterologous DGA1 polynucleotides encoding DGA1 proteins, or functionally active portions thereof, having the capability of producing increased lipid production and possessing the characteristic of enhanced glucose consumption efficiency.

Abstract: The invention relates to: a method for producing a protease in yeast, comprising providing a yeast host cell comprising at least one polynucleotide expression construct encoding a first polypeptide secreted by the host cell in translational fusion with a second polypeptide, wherein the second polypeptide is a protease having a mature amino acid sequence similar to that of SEQ ID NO:2; the corresponding yeast host cell and the expression construct.

Abstract: An expression vector which is capable of overexpressing a protein of interest in a host cell, a host cell comprising the expression vector, and a method of producing a protein of interest are provided.

Abstract: The invention relates to a method for the prevention or reduction of haze in a beverage by the addition of a prolyl-specific and/or alamine specific endoprotease and the new beverages obtainable by the method according to the invention. It also relates to new endoproteases. It also relates to methods as described above wherein auxiliary enzymes are used in combination with the specific endoprotease. Sequence information of a genomic DNA, cDNA as well as protein sequences are provided.

Abstract: The invention relates to a method for the prevention or reduction of haze in a beverage by the addition of an prolyl-specific endoprotease and to new beverages obtainable by the method according to the invention. It also relates to new endoproteases. Sequence information of a genomic DNA, cDNA as well as protein sequences.

Abstract: A method of enhancing heterologous protein secretion in a yeast cell is disclosed. In one embodiment, the method comprising the steps of engineering a yeast cell to overexpress at last one gene selected from the group consisting of CCW12, CWP2, SED1, RPP0, ERO1 and their homologs, supplying the yeast cell with a nucleic acid encoding a heterologous protein and obtaining increased expression of the heterologous protein, wherein the expression is increased relative to the protein expression in a yeast cell that does not overexpress a gene selected from the group consisting of CCW12, CWP2, SED1, RPP0, ERO1 and their homologs.

Abstract: A method of detection of cells, microorganisms, or molecules by the use of various combinations of fluorogens and a chromogens which yield fluorophores and chromophores when cleaved by specific enzymes and which can be viewed by UV and visible light. Included is the method of application of a family of compounds producing both insoluble fluorophores and chromophores identified as dual enzyme substrates.

Abstract: The present invention discloses manufacturing method of low temperature protease and special yeast strain, which can generate low temperature protease in the condition of low temperatures. More particularly, the present invention is to obtain a protease (preferably Cold-adapted protease PI12) using the marine strain of the Leucosporidium antarcticum sp. (NCYC accession no: 3391) for possible use in industries.

Abstract: Two genes which encode polypeptides that mediate post-prenylation processing steps in CAAX polypeptides such as Ras are provided. The two genes (AFC1 and RCE1) encode polypeptides that mediate the removal of the AAX tripeptide from the CAAX polypeptide following prenylation. The genes and encoded polypeptides provide assays for testing compounds for an effect on post-prenylation processing steps. A heat shock assay for assessing Ras activity is also provided.

Abstract: The present invention is directed to expression vectors comprising a polynucleotide that encodes a human telomerase reverse transcriptase (hTRT) protein, variant, or fragment. The present invention is also directed to host cells that comprise expression vectors comprising a polynucleotide that encodes a hTRT protein variant, or fragment.

Abstract: The present invention relates to a novel methods and compositions for producing hyper and hypo allergenic compositions. Specifically, the present invention comprises neutralizing or reducing the ability of T-cells to recognize epitopes and thus prevent sensitization of an individual to the protein. Alternatively, T-cell epitopes are mutated to produce increased immunogenic reactions.

Abstract: An improved infant formula resulting in reduced constipation, abdominal discomfort and gastrointestinal problems, comprises at least one protein component having a phosphorus content of less than 0.75 g P/100 g protein, and at least one lipid component that can be easily digested by an infant. Preferably, it further comprises at least one prebiotic component, and at least one viscosity-improving component. The protein fraction of the formula is preferably a hydrolysate prepared by hydrolysing a protein starting material, especially a whey protein with a combination of at least one endo- and at least one exoproteinase.

Abstract: The present invention provides a process for economically producing N-acetylneuraminic acid without using expensive materials such as pyruvic acid and phosphoenolpyruvic acid. The process comprises: allowing (i) a culture of a microorganism having N-acetylneuraminic acid aldolase activity or N-acetylneuraminic acid synthetase activity, or a treated matter of the culture, (ii) a culture of a microorganism capable of producing pyruvic acid or a treated matter of the culture, or a culture of a microorganism capable of producing phosphoenolpyruvic acid or a treated matter of the culture, (iii) N-acetylmannosamine, and (iv) an energy source which is necessary for the formation of pyruvic acid or phosphoenolpyruvic acid to be present in an aqueous medium to form and accumulate N-acetylneuraminic acid in the aqueous medium; and recovering N-acetylneuraminic acid from the aqueous medium.

Abstract: The present invention relates to the use of a novel enzyme and its encoding gene for transformation. More specifically, the invention relates to the use of a gene encoding an enzyme with acyl-CoA:diacylglycerol acyltransferase activity. This gene expressed alone in transgenic organisms will increase the total amount of oil (i.e. triacylglycerols) that is produced.

Abstract: A novel enzyme which has an activity to release side chain carboxyl groups and ammonia from a protein by acting upon side chain amido groups in the protein. This invention relates to a method for the production of an enzyme, which comprises culturing in a medium a strain that belongs to a bacterium classified into Cytophagales or Actinomycetes and has the ability to produce an enzyme having a property to deamidate amido groups in protein, thereby effecting production of said enzyme, and subsequently collecting said enzyme from the culture mixture. It also relates to a method for the modification of protein making use of a novel enzyme which directly acts upon amido groups in protein as well as to an enzyme which has a property to deamidate amido groups in protein and a gene which encodes said enzyme.

Abstract: Disclosed is a method of producing a compound with &bgr;1-4 linkage which contains the lactosamine structure involving reacting at least one donor substance Gal&bgr;OR where R is an organic group, and at least one acceptor substance which is a glucopyranosamino derivative having the formula GlcNR″—R′″, wherein NR″ is an azido, 2-N-acetyl-, 2-N-phtalimido, or an organic group bound to the 2-N-group of glucosamine, wherein R′″ is a glycosidically bound fluoro or is an O-, C-, N- or S-glycosidically bound aliphatic or aromatic compound, with the proviso that if NR″ is NHAc then R′″ is not OH and if NR″ is not NHAc then R′″ may be OH, in the presence of Bullera singularis or an E.C. group 3.2 glycosidase of essentially the same structure as an E.C. Group 3.

Abstract: The present invention relates to a novel improved protein mutant which produces low allergenic response in humans compared to the parent of that mutant. Specifically, the present invention comprises neutralizing or reducing the ability of T-cells to recognize epitopes and thus prevent sensitization of an individual to the protein.

Abstract: An object of the present invention is to provide novel genes and gene group involved in cellulose synthesis of microorganisms. The present invention relates to a gene group encoding cellulase, cellulose synthase complex, &bgr;-glucosidase and the like, and to novel &bgr;-glucosidase.

Abstract: The present invention discloses a method for culturing microorganisms having a methanol metabolic pathway in which an expression unit is introduced that comprises a target gene linked downstream from a promoter that can be induced by methanol; wherein, during the culturing period, and including the period during which methanol is continuously or periodically added, the rate of addition is adjusted to a rate equal to or less than the maximum methanol consumption rate of said microorganisms.

Abstract: The present invention relates to a process for producing optically active 3-quinuclidinol or derivatives, wherein a racemic 3-quinuclidinol ester represented by the general formula (I): ##STR1## wherein R represents a straight-chain or branched alkyl group, and (H.sup.+) represents that said ester may be in the form of a salt formed with a mineral acid or an organic acid, is reacted with a microorganism belonging to the genus Aspergillus, Rhizopus, Candida or Pseudomonas having the ability to asymmetrically hydrolyze said ester linkage, a culture of said microorganism, a treated material from said microorganism, an enzyme produced by said microorganism, or an enzyme derived from swine or cattle.According to the present invention, there is provided a process for easily producing optically active 3-quinuclidinol derivatives which are important synthetic intermediates for pharmaceutical preparations etc.

Abstract: A process for the production of a desired polypeptide comprising the steps of: (1) transforming host cells with an expression vector comprising a gene coding for a fusion protein comprising a desired polypeptide and a protective polypeptide; (2) culturing the transformed host cells so as to express said gene to produce a fusion protein; and (3) excising the desired polypeptide from the fusion protein with a protease intrinsic to the host cells. According to the present invention, a large amount of a desired polypeptide can be produced at a low cost. Especially according to the present invention, a large amount of S. aureus V8 protease can be efficiently produced at low cost using a safe host such as E. coli according to gene recombination procedures.

Abstract: An isolated and purified enzyme exhibiting protease activity at a pH of 4-7 which exhibits protease activity in 5% hydrogen peroxide and which is encoded by a DNA sequence which hybridizes to a DNA sequence depicted in SEQ ID NO. 1 or 2.

Abstract: A method for transamidating a peptide substrate having a P.sub.1 amino acid residue with a positively charged side chain. According to the invention, carboxypeptidase Y is modified to substitute at least one amino acid having a negatively charged side chain in an S.sub.1 subsite. Additionally, the modified carboxypeptidase Y can include substituted amino acid residues in an S.sub.1 ', S.sub.2 and/or S.sub.3 subsite to accommodate a specific peptide substrate.

Abstract: The present invention relates to methods of producing a polypeptide, comprising: (a) introducing into a respiratory-defective mutant of a cell (i) one or more first nucleic acid sequences which complement the respiratory defect and (ii) a second nucleic acid sequence which encodes the polypeptide; (b) cultivating the cell containing the first and second nucleic acid sequences in a culture medium under aerobic conditions suitable for expression of the first and second nucleic acid sequences; and (c) isolating the polypeptide from the cultivation medium of the cell. The present invention also relates to methods for disrupting a gene in a respiratory-deficient mutant cell. The present invention further relates to respiratory-deficient mutant cells and methods for obtaining such mutant cells.

Abstract: Disclosed are methods that can be used to (1) measure the level of polysaccharide in a sample; (2) measure the ability of a compound to degrade a polysaccharide; (3) measure the ability of a compound to modulate polysaccharide synthesis; and (4) identify or distinguish a polysaccharide, and hence organism, for diagnostic purposes in clinical medicine or research. The invention stems from Applicant's discovery that polysaccharides have multiple binding sites for polysaccharide binding moieties (PBM, e.g., wheat germ agglutinin (WGA)). In each method, one PBM links the polysaccharide to a substrate, and a tagged PBM is used to detect the polysaccharide. All of these methods can be carried out rapidly and quickly in the wells of a microtiter plate, thus permitting high through-put screening of samples or test compounds.

Abstract: Kex2 protease derivatives obtained by transforming methanol-assimilating with expression vectors containing DNA coding for any amino acid sequence which is an amino acid sequence of Kex2 protease wherein the N-terminus is the Met at position 1 and the C-terminus is one of the amino acids between positions 618 (inclusive) and 698 (inclusive), or a modification of that amino acid sequence, culturing the resulting transformants and recovering the derivatives from the cultures, as well as gene systems coding for the derivatives and a method for producing the Kex2 protease derivatives using the gene systems. Also, a method for excision of desired peptides using the Kex2 protease derivatives.

Abstract: A DNA construct comprising a DNA sequence encoding an enzyme exhibiting protease activity, which DNA sequence comprises the DNA sequence shown in SEQ ID No. 1 or 2 or an analog of any of these sequences being at least 80% homologous to the DNA sequence shown in SEQ ID No. 1 or 2. The proteases encoded by the DNA sequences have an acid pH optimum.

Abstract: The present invention relates to a novel metalloprotease obtainable from a fungus having increased proteolytic activity. Additionally, the invention related to isolated nucleic acid fragments encoding said metalloprotease as well as vectors, DNA constructs, and recombinant host cells comprising said nucleic acid fragments.

Abstract: The present invention relates to a novel metalloprotease obtainable from a fungus having increased proteolytic activity. Additionally, the invention related to isolated nucleic acid fragments encoding said metalloprotease as well as vectors, DNA constructs, and recombinant host cells comprising said nucleic acid fragments.

Abstract: The invention relates to serine protease variants derived from precursor serine proteases via recombinant and/or chemical methods to form protease variants having improved peptide ligase activity. The invention also includes novel ligation substrates which in combination with the serine protease variants and a second ligation substrate are capable of forming a ligation product. The invention also relates to methods for forming such ligation products and the products formed thereby.

Abstract: The present invention relates to a novel metalloprotease obtainable from a fungus having increased proteolytic activity. Additionally, the invention related to isolated nucleic acid fragments encoding said metalloprotease as well as vectors, DNA constructs, and recombinant host cells comprising said nucleic acid fragments.

Abstract: A laundry detergent for washing fabrics composed of proteinogenic fibers is comprised of at least one surfactant and a proteolytically active amount of a protease having a keratinase/caseinase activity ratio of less than about 0.80.

Abstract: The invention herein provides bleaching compositions comprising a protease enzyme which is a carbonyl hydrolase variant having an amino acid sequence not found in nature, which is derived by replacement of a plurality of amino acid residues of a precursor carbonyl hydrolase with different amino acids, wherein said plurality of amino acid residues replaced in the precursor enzyme correspond to position +76 in combination with one or more of the following residues: +99, +101, +103, +104, +107, +123, +27, +105, +109, +126, +128, +135, +156, +166, +195, +197, +204, +206, +210, +216, +217, +218, +222, +260, +265, and/or +274, where the numbered positions corresponds to naturally-occurring subtilisin from Bacillus amyloliquefaciens or to equivalent amino acid residues in other carbonyl hydrolases or subtilisins (such as Bacillus lentus subtilisin) and a bleaching agent.

Abstract: The present invention relates to novel yeast strains which produce a heterologous precursor protein having a dibasic amino acid processing site which can be processed into at least one cleavage protein by a dibasic amino acid processing endoprotease. Such novel yeast strains are useful for identifying compounds capable of inhibiting infectious agents, such as viruses, that depend upon dibasic amino acid processing endoprotease cleavage for effective propagation and/or infectivity.

Abstract: The present invention is directed to a method for screening samples for the identification of agents exhibiting potential fungicidal and insecticidal activity for a wide variety of agricultural, medical and pharmaceutical uses. The method utilizes cells that comprise a plasmid-born CTS gene of Saccharomyces cerevisiae, which allows for over expression of chitinase.

Abstract: The isolation and characterization of genes involved in proteolytic processing in species of the genus Pichia is described. The availability of such genes has enabled the generation of strains of Pichia which are deficient in proteolytic activity, which strains are useful as hosts for the expression of proteolytically sensitive recombinant products. The isolation and characterization of additional genes from species of the genus Pichia is also described, as well as uses therefore.

Abstract: Nonaqueous gelled automatic dishwashing compositions containing a mixture of a protease enzyme and an amylase enzyme have been found to be very useful in the removal of protein and carbohydrate soils from dishware at operating temperatures of 100.degree. F. to 140.degree. F.

Abstract: The present invention relates to an improved system for producing mature heterologous proteins, and especially hirudin, by means of yeasts comprising an expression vector containing a sequence coding for the heterologous protein. This improvement is characterized by amplification of the KEX2 gene of yeast, coding for the endoprotease yscF. The amplification is carried out either by integration of one or more copies of all or part of the KEX2 gene in the yeast genome, or by insertion of one or more copies of all or part of the KEX2 gene into the vector for expression of the heterologous protein.

Abstract: A high density, enzyme-containing powder detergent composition including a combination of alkaline proteases for improved cleaning characteristics and a combination of different density sodium carbonates for improved dispensing characteristics.

Abstract: The present invention provides a human polyfuctional protease chracterized in that the protease has unique enzymological and physicochemical properties:

Abstract: The present invention relates to a novel method to identify compounds that inhibit proteolytic cleavage by dibasic amino acid processing endoproteases that includes contacting a yeast strain with a putative inhibitory compound under conditions in which, in the absence of the compound, the yeast strain can cleave a precursor protein having a dibasic amino acid processing site and determining if the putative inhibitory compound inhibits cleavage of the precursor protein. The present invention includes a method to identify compounds capable of inhibiting infectious agents, such as viruses, that depend upon dibasic amino acid processing endoprotease cleavage for propagation. The present invention also includes assay kits based on such a method.

Abstract: A ubiquitin-specific protease which cleaves ubiquitin from any protein or peptide to which ubiquitin is joined and the gene encoding the protease are disclosed. The protease specifically cleaves the peptide bond in a fusion of ubiquitin to a protein or peptide between the carboxyl-terminal amino acid residue of a ubiquitin moiety and the .alpha.-amino group of any non-ubiquitin protein or peptide to which ubiquitin is joined. Recombinant expression vectors containing a DNA sequence encoding the ubiquitin-specific protease can be used to transform cells for production of the protease or to provide the cell with the ability to proteolytically deubiquitinate, in vivo, ubiquitin fusions co-produced by the cell. The protease can also be isolated and used to deubiquitinate ubiquitin fusions in vitro.

Abstract: This invention is a method to rupture microbial cells in order to recover intracellular material in the cells comprising:a) treating the cells with carbon dioxide under pressure sufficient to enter the cells for time sufficient to allow enough carbon dioxide into the cells to effect later rupture, thenb) suddenly releasing the applied fluid pressure on the cells so that the outer wall or membrane of the cells is ruptured by the expansion of carbon dioxide within the cell. Preferably the remaining intracellular material of the cells is separated and recovered. Also the treatment can be in conjunction with lytic enzyme to increase rupture rates. Preferably the enzyme remains active and protein in the cells retains its native state in the ruptured cell suspension. The preferred time for treating is from between about one hour and about fifteen hours. It is also preferred to treat initially at a pressure of from above about 500 psi gage to about 5000 psi and a temperature of about 10.degree. to about 85.degree.

Abstract: A ubiquitin hydrolase is provided having a purity of at least 70% homogeneity based on the weight of the total protein in the composition. Also provided are DNA sequences encoding ubiquitin hydrolases, as well as expression systems for their recombinant production. Processes are provided for purification of a ubiquitin hydrolase from eukaryotes and for its use in recovering any desired polypeptide free from its fusion at its N-terminus with ubiquitin.

Abstract: Alkaline protease which has leucine or isoleucine in the place of valine at the amino acid number 40 of wild type alkaline protease, a gene encoding the amino acid sequence of alkaline protease which has the substitution as described above, a recombinant DNA comprising said gene, a method of producing the above described alkaline protease, and a DNA fragment used for the expression of a gene.

Abstract: A KEX2 endoprotease produced by a recombinant DNA technique, a KEX2 endoprotease shortened at the C-terminal of a native enzyme and still containing a C-terminal hydrophobic region, a soluble KEX2 endoprotease without a C-terminal hydrophobic region; DNA's coding for the above-mentioned enzymes; expression plasmids containing the DNA; hosts transformed with the plasmid; a process for production of the above-mentioned enzymes using the transformed host; and a process for production of biologically active polypeptide using the above-mentioned enzyme.

Abstract: A KEX2 endoprotease produced by a recombinant DNA technique, a KEX2 endoprotease shortened at the C-terminal of a native enzyme and still containing a C-terminal hydrophobic region, a soluble KEX2 endoprotease without a C-terminal hydrophobic region; DNA's coding for the above-mentioned enzymes; expression plasmids containing the DNA; hosts transformed with the plasmid; a process for production of the above-mentioned enzymes using the transformed host; and a process for production of biologically active polypeptide using the above-mentioned enzyme.

Abstract: Ubiquitin hydrolase is provided having a purity of at least 70% homogeneity based on the weight of the total protein in the composition, which hydrolase hydrolyzes a ubiquitin-polypeptide conjugate at the amide bond linking the ubiquitin and polypeptide, thereby yielding intact polypeptide with an unconjugated, mature N-terminus. Also provided are DNA sequences encoding such ubiquitin hydrolase, as well as expression systems for its recombinant production. Processes are provided for purification of ubiquitin hydrolase from eukaryotes and for its use in recovering any desired polypeptide free from its fusion at its N-terminus with ubiquitin.

Abstract: Method for producing lignin-peroxydase from the fungus Phanerochaete chrysosporium. The method comprises a first step of culture wherein the phospholipids and the emulsified fatty acids are added to the culture medium; a second step during which veratrylic alcohol is added to the culture medium, the culture medium being partially renewed for the second culture step and totally renewed for the third and fourth culture steps, by varying the content of constituents in said medium. Application to the production of lignin-peroxydase with important yields.