Abstract: The invention includes novel fusion DNA sequences encoding fusion polypeptides which when expressed in a filamentous fungus result in the expression of fusion polypeptides which when secreted result in increased levels of secretion of the desired polypeptide as compared to the expression and secretion of such polypeptides from filamentous fungi transformed with previously used DNA sequences. The fusion DNA sequences comprise from the 5′ terminus four DNA sequences which encode a fusion polypeptide comprising, from the amino to carbonyl-terminus, first, second, third and fourth amino acid sequences. The first DNA sequence encodes a signal peptide functional as a secretory sequence in a first filamentous fungus. The second DNA sequence encodes a secreted polypeptide or portion thereof which is normally secreted from the same filamentous fungus or a second filamentous fungus. The third DNA sequence encodes a cleavable linker polypeptide while the fourth DNA sequence encodes a desired polypeptide.

Abstract: There are described methods for making mutant subtilisins, the methods comprising obtaining a DNA fragment from a Bacillus subtilisin and introducing a mutation into the fragment by substituting at least one amino acid, transforming a suitable host cell with the mutated DNA, recovering a mutant subtilisin and screening the mutant subtilisin for certain altered enzymatic properties.

Abstract: Processes for producing various heterologous polypeptides which when expressed are either incorrectly processed and hence asssociated with the surface of the host cell or are not processed to mature form. More specifically, processes for the production of heterologous non-human carbonyl hydrolases expressed either in host cells incapable of producing enzymatically active endoprotease or host cells deficient in enzymatically active extracellular endoprotease are disclosed. Such non-human carbonyl hydrolases generally are incapable of autoproteolytic maturation and become associated with the surface of expression hosts which are deficient in enzymatically active extracellular endoprotease. Processes for preparing non-human carbonyl hydrolase and heterologous polypeptides which are expressed as part of a fusion polypeptide are also disclosed, as well as non-human carbonyl hydrolases which are substantially free of the host cell membrane with which they are normally associated.

Abstract: A substantially enzymatically pure hydrolase is provided which is secreted by and isolatable from Pseudomonas mendocina ATCC 53552. Cloning the gene expressing the hydrolase into a suitable expression vector and culturing, such as fermenting the E. coli strain JM101 harboring a plasmid designated pSNtacII, has been found to provide surprisingly high yields of the hydrolase.

Abstract: This invention provides a method for killing fungal cells without lysing in fermentation processes in order to prepare the fermentation mixture for processing to recover or extract an extracellularly expressed enzyme from the fermentation mixture. A preferred method of this invention comprises adjusting the pH of the fermentation mixture to less than 2.79 using a mineral acid, then adding sufficient acetic acid to the mixture to affect a substantially complete cell kill in mixture. A salt of the acetic acid can be used. The organic acid or salt can be added, then the pH adjusted to the desired level. Other organic acids can be used, in which case the pH of the mixture is adjusted to the pK.sub.a of the selected organic acid before the organic acid is added to the mixture.

Abstract: There are provided methods for making a mutant Bacillus subtilisin having altered oxidative stability, the methods comprising obtaining DNA fragment consisting of a region coding for a Bacillus subtilisin, and introducing a mutation into said DNA fragment such that the mutation is introduced in a region encoding a methionine, tryptophan, cysteine or lysine, sensitive to oxidation, such that upon expression of the mutant subtilisin one or more codon regions encoding for methionine, tryptophan, cysteine or lysine is replaced with an amino acid other than methionine, tryptophan, cysteine or lysine, preferably alanine or serine.

Abstract: There are described certain DNA sequences which encode subtilisins wherein the amino acid sequence of such substilisins has been modified at a position equivalent to +225 in Bacillus amyloliquefaciens, such that an amino acid selected from the group consisting of alanine, leucine, methionine, glutamine, valine and serine, has been substituted for the amino acid residues naturally occuring at such position.

Abstract: Recombinant host bacteria and plasmids for making the bacteria using recombinant DNA techniques are described. The plasmids contain DNA coding for subtilisin with an amino acid substitution. Expression of the plasmid DNA results in production of a modified subtilisin.

Abstract: There are described, normally sporulating mutant Bacillus strain(s) which produce no detectable proteolytic activity during any phase of its growth. The absence of detectable proteolytic activity is due to the deletion of one or more codons specifying the mature subtilisin protease and the mature neutral protease. Also described are methods for producing such normally sporulating, protease deficient Bacillus mutants.

Abstract: There are described certain subtilisins wherein the amino acid sequence of such subtilisins has been modified at a position equivalent to +225 in Bacillus amyloliquefaciens, such that an amino acid selected from the group consisting of alanine, leucine, methionine, glutamine, valine, and serine, has been substituted for the amino acid residue naturally occurring at such position.

Abstract: This invention relates to cleaning compositions and methods for using them. Particularly, the invention relates to compositions comprising a surfactant and a cutinase enzyme. A preferred cutinase is derived from Pseudomonas putida ATCC 53552. Excellent cleaning is obtained with a surfactant mixture containing sodium dodecyl sulfate and octoxynol.

Abstract: The invention relates to a novel method for preferentially entraining a lipase or protease substantially into cheese curds when making curds and whey. An insoluble enzyme particle size of at least about 0.20 microns is selected or formed, and upon curd formation the insoluble particle will substantially be entrained therein, thus, leading to a substantial reduction in the amount of enzyme lost in the whey without the need to add additional complexing or immobilizing agents.

Abstract: The invention relates to a method of milling grain, especially corn, comprising cleaning the grain, steeping the grain in water to soften it, and then milling the grain with a cellulase enzyme.

Abstract: A cloned subtilisin gene has been modified at specific sites to cause amino acid substitutions at certain spots in the enzyme. The modified enzyme, preferably produced by Bacillus, is useful in combination with detergents.

Abstract: Contact lens cleaning compositions and a method which comprises treating soft contact lenses with general purpose proteases in combination with endoproteinase lys-C are effective in dissolving away and hydrolyzing lysozyme, the major protein component of tears.

Abstract: The present invention relates to a novel lipolytic enzyme derived from a novel Aspergillus microorganism. Cheese aged in the presence of a low concentration of this lipolytic enzymes ripens faster than with conventional lipolytic enzymes and without any associated rancidity.

Abstract: This application represents an invention based upon the discovery of a novel thermophilic organism, its isolation from natural source and to the unique thermostable hydrolytic activity produced by this bacterium, this activity being useful for industrial purposes.

Abstract: An improved process for removing insoluble nitrogen-containing compounds from cured tobacco uses alkali or a combination of protease and nonprotease depolymerase, rather than simple protease extraction.The method of the invention is more efficient and results in a more effective extraction of protein.

Abstract: The present invention relates to a novel lipolytic enzyme derived from a novel Aspergillus microorganism. Cheese aged in the presence of a low concentration of this lipolytic enzymes ripens faster than with conventional lipolytic enzymes and without any associated rancidity.

Abstract: A cloned subtilsin gene has been modified at specific sites to cause amino acid substitutions at certain spots in the enzyme. The modified enzyme, preferably produced by Bacillus, is useful in combination with detergents.