Abstract: New mutant glucose isomerases are provided exhibiting improved properties under application conditions. These glucose isomerases are obtained by expression of a gene encoding said enzyme, having an amino acid sequence which differs at least in one amino acid from the wildtype glucose isomerase. Preferred mutant enzymes are those derived from Actinoplanes missouriensis glucose isomerase.

Abstract: A method for selecting amino acid residues is disclosed which upon replacement will give rise to an enzyme with an altered pH optimum. The method is specific for metalloenzymes which are inactivated at low pH due to the dissociation of the metal ions. The method is based on altering the pK.sub.a of the metal coordinating ligands or altering the K.sub.ass for the metal binding. New glucose isomerases with an altered pH optimum are provided according to this method. These altered properties enable starch degradation to be performed at lower pH values.

Abstract: A method for selecting amino acid residues is disclosed which upon replacement will give rise to an enzyme with an altered substrate specificity. New mutant glucose isomerases with an altered substrate specificity are provided according to this method. These altered properties are useful in starch degradation and in other sugar conversion reactions.

Abstract: A genetically engineered glucose isomerase with improved affinity for D-glucose and the method of preparation of such a glucose isomerase are disclosed. The glucose isomerase is obtained by mutagenizing the gene of a naturally occurring glucose isomerase such that a smaller amino acid replaces a larger amino acid in the catalytic site. In an especially advantageous embodiment of the present invention, the Clostridium glucose isomerase sequence is mutated and the residue replaced with a smaller amino acid is either Trp.sub.139 or Val.sub.186.

Abstract: Process for manufacture of D-xylose characterized by the fact that:in a first step, syrup of D-xylulose is subjected to an enzymatic isomerization in M.sub.3 providing a mixture of D-xylose and D-xylulose,in a second step, the abovesaid mixture is subjected to chromatographic treatment in M.sub.4 leading to at least two fractions of which one is highly enriched in D-xylose (fraction X.sub.1) et of which the other is highly enriched in D-xylulose (fraction X.sub.2),in a third step, the fraction X.sub.2 is recycled through a pipe P to M.sub.3,the D-xylose being recovered from the fraction X.sub.1, the latter can also be subjected directly to a hydrogenation step.

Abstract: This invention is in the field of glucose isomerization enzymes. More specifically, the invention is directed to a novel xylose isomerase, a process for the preparation of this enzyme, the use of this enzyme in glucose isomerization processes, and glucose isomerization processes. The enzyme is preferably derived from Thermotoga maritima or Thermotoga neapolitana. The enzyme has a temperature optimum above 90.degree. C., pH optimum in the range of from 6 to 7 and a residual activity at 90.degree. C. of more than 40% after 30 minutes and/or residual activity at 98.degree. C. of more than 20% after 30 minutes. The enzyme can also be in immobilized form.

Abstract: This invention provides a method for the maintenance of continuous activity of an immobilized enzyme reaction by initially underloading an adsorbent with a desired enzyme, monitoring the activity of the enzyme and periodically adding fresh enzyme to maintain a constant, continuous level of activity until the maximum carrying capacity of the support is reached. The method is preferably carried out when continuously isomerizing glucose to fructose with glucose isomerase adsorbed to a weakly basic anion exchange resin.

Abstract: In a process for growing enzyme crystals, small crystals are continuously removed from a crystallizer, dissolved and returned to the crystallizer to maintain a supersaturated state. The method permits the growing of large crystalline enzymes of uniform size of about 0.5 to 1 mm. Solid materials can be coated with crystalline enzymes by placing a solid material in the crystallizer such that crystals deposit on the solid material. The process is preferably used to produce crystalline glucose isomerase.

Abstract: Xylose isomerase (XI) muteins useful in the conversion of glucose to fructose or xylose to xylulose are obtained in usable amounts by protein structural and recombinant DNA methods, including x-ray crystallography, cloning, computer graphic modeling and site-directed mutagenesis and expression of the bacterial DNA sequences encoding native procaryotic xylose isomerase. These native sequences are altered to encode the xylose isomerase muteins having improved catalytic function and/or thermostability.

Abstract: A composite containing DEAE-cellulose agglomerated with a hydrophobic polymer is treated with tap water, deionized water or a dilute salt solution at a temperature of at least 60.degree. C. for a time sufficient to increase adsorption capacity for charged macromolecules by at least about 30%. Treatment is preferably at a temperature of about 80.degree. to about 100.degree. C. for a time period of about one-half hour to about 5 hours. The macromolecule is preferably a protein such as the enzyme, glucose isomerase.

Abstract: The subject invention concerns a process for stabilizing intact or ruptured microbial cells having glucose isomerase associated therewith. Specifically exemplified is a process for stabilizing glucose isomerase producing cells of a microorganism belonging to the genus Ampullariella. In the invention process the whole or ruptured microbial cells are contacted with a partially carboxyalkylated- or partially phosphonoalkylated-cationic polyelectrolyte, for example, a partially carboxymethylated polyethyleneimine to flocculate and stabilize the cells. The flocculated cells are further stabilized by encapsulation with a partially carboxyalkylated- or partially phosphonoalkylated-cationic polyelectrolyte. The encapsulation can be done prior to or after the flocculated cells are crosslinked. The net effect is manifested by a dramatic increase in the half-life of the glucose isomerase.

Abstract: Polysulfonium salts that can react with nucleophilic groups and covalently cross-link are used to immobilize enzymes or enzyme-containing cellular material. Some of the polysulfonium salts can both flocculate and covalently cross-link. Replacement of the cross-linker, glutaraldehyde, with the polysulfonium salt results in greater retention of enzyme activity during immobilization. Immobilization is carried out by forming a mixture of an enzyme or enzyme-containing cellular material and the polysulfonium salt and subjecting the mixture to conditions such that sulfonium ions react with nucleophilic groups contained by the enzyme or cellular material to form a covalently cross-linked and water insoluble product. The enzyme or cellular material may be flocculated with a flocculating agent prior to cross-linking with the polysulfonium salt. The polysulfonium salt can be a polymer containing sulfonium groups.

Abstract: A glucose isomerase useful for the conversion of glucose to fructose can be prepared by growing under aerobic conditions a culture of a species of Microbacterium in a medium containing appropriate nutrients and then recovering the enzyme.

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 method comprising stimulating the biosynthesis of glutathione in mammalian cells by contacting the cells with an effective amount of a compound of the formula: ##STR1## wherein R is a (CHOH).sub.n CH.sub.2 OH and wherein n is 1-5, preferably R is derived from a D-aldose monosaccharide.

Abstract: Xylose isomerase, from strains of the Streptomyces murinus cluster, a method for production of such xylose isomerase, immobilized xylose isomerase and a method for isomerization of glucose to fructose therewith.

Abstract: The disclosure is directed to the resolubilization of an insoluble glucose isomerase-amine complex, wherein the amine has the general formula: ##STR1## The insoluble enzyme complex may be resolubilized to produce a stable concentrated and purified glucose isomerase preparation by reaction with a resolubilizing mixture comprising a cation exchange resin.

Abstract: The disclosure is directed to the treatment of a glucose isomerase extract with an amine having the general formula: ##STR1## An insoluble complex containing enzyme activity is formed. The insoluble enzyme complex may be resolubilized to produce a stable concentrated and purified glucose isomerase preparation.

Abstract: This invention relates to a process for the production of a purified glucose isomerase enzyme which comprises contacting an enzyme extract containing glucose isomerase and impurities with a first polysulfone membrane not normally permeable to glucose isomerase, in the presence of a salt concentration capable of selectively inducing permeation of glucose isomerase through the membrane, and obtaining a glucose isomerase containing permeate.

Abstract: The present invention relates to a process for the production of a purified glucose isomerase which comprises contacting an impure extract containing glucose isomerase and soluble impurities with a weakly basic ion exchange material known to adsorb glucose isomerase; adding a first salt solution of a concentration which removes unadsorbed and weakly adsorbed impurities, but not glucose isomerase; and adding a second, buffered salt solution which elutes the adsorbed glucose isomerase.

Abstract: A process is disclosed for chemically modifying naturally occurring proteins to produce enzyme-like modified proteins. The process comprises partially denaturing a cofactor containing holoprotein by removal of the cofactor to produce a partially denatured cofactorless or so-called apoprotein. The partially denatured protein is contacted with an inhibitor of a selected model enzyme and cross-linked. The resultant protein product is an enzyme-like modified protein having the catalytic characteristics of the model enzyme whose inhibitor is contacted with the partially denatured apoprotein.

Abstract: Process for preparing fructose by treating starch with alpha-amylase, contacting the resulting liquefied starch with glucoamylase to hydrolyzed said starch to glucose, and isomerizing at least part of the resulting glucose to fructose by contacting said glucose with glucose isomerase. The three enzymes are obtained from organisms of the Basidiomycetes class of fungi.

Abstract: The invention relates to a method for producing a glucose isomerizing enzyme which is functional at temperatures over 90.degree., utilizing a strain microorganisms having the identifying characteristics of Arthrobacter sp. ATCC 21748.

Abstract: Disclosed is a process for purifying an enzyme contained in a solution such as cell extract liquor or fermentation culture liquor. The crude enzyme solution is brought into contact with either a strongly acidic cation exchange resin of high porous type or a strongly basic anion exchange resin of high porous type to adsorb the enzyme on the resin. An eluting agent is then passed through the resin to elute out the enzyme as a purified enzyme solution.

Abstract: A mutant Streptomyces thermoviolaceus, NRRL 15615, elaborates a thermostable glucose isomerase in high yield, constitutively. The GI produced shows a negligible loss in activity when heated at 90.degree. C. in high fructose corn syrup for 5 minutes relative to the same treatment at 80.degree. C. The mutant microorganism produces about 1500 units of glucose isomerase per gram of dry weight cells in the absence of xylose.

Abstract: A mutant Streptomyces coelicolor, NRRL 15398, produces glucose isomerase constitutively at a level at least as great as the parent does inductively in common growth media. The isomerase can be effectively used to isomerize glucose to fructose in a continuous process using immobilized enzyme.

Abstract: Disclosed is a method of increasing the production of enzymes involved in carbohydrate metabolism by bacterium of the species F. arborescens or E. coli. The method involves growing the bacterium on an aqueous nutrient medium containing glucose oxime.

Abstract: A high activity immobilized enzyme composite is prepared by covalently bonding a second enzyme layer to a first enzyme layer that is immobilized to a carrier. The carrier is preferably silica gel which has been activated by treatment with a strong base followed by treatment with a strong acid. The first enzyme layer is covalently bonded to the activated silica gel with an aminosilane and a polyfunctional reactant, and the second enzyme layer is covalently bonded to the first layer with a polyfunctional reactant. Third, fourth and more successive enzyme layers may be covalently bonded. The composite has high activity per unit volume, superior stability and good half-life.

Abstract: Process for preparing L-fructose from L-glucose by contacting L-glucose with xylose isomerase produced by a microorganism of the genus Candida.

Abstract: Acidophilic and acidoduric streptomycetes strains have been found to produce carbohydrases. These Streptomyces effectively elaborate glucose isomerase under acid conditions typically unfavorable for growth of conventional glucose isomerase producing Streptomyces. Sterilization of the culture and production media may be avoided by selectively propagating newly discovered Streptomyces acidodurans under acidic conditions which will effectively eliminate contaminating micro-organisms. The Streptomyces acidodurans herein also have the ability to undergo cultivation and elaborate glucose isomerase over a relatively broad pH range. Constitutive streptomycetes strains have also been isolated. Glucose isomerases derived from these Streptomyces strains are particularly effective for isomerizing glucose syrups to fructose-containing syrups.

Abstract: Process for preparing fructose by treating starch with alpha-amylase, contacting the resulting liquefied starch with glucoamylase to hydrolyze said starch to glucose, and isomerizing at least part of the resulting glucose to fructose by contacting said glucose with glucose isomerase. The three enzymes are obtained from an organism of the Basidiomycetes class of fungi.

Abstract: Glucose isomerase is produced by cultivating fungi of the Basidiomycetes class and is used for isomerizing glucose to fructose.

Abstract: Process for preparing fructose from liquefied starch by contacting the liquefied starch with glucoamylase to hydrolyze said starch to glucose and contacting the glucose so produced with glucose isomerase to isomerize at least a part of the glucose to fructose. The glucoamylase and glucose isomerase are obtained from an organism of the Basidiomycetes class of fungi.

Abstract: Acidophilic and acidoduric streptomycetes strains have been found to produce carbohydrases. These Streptomyces effectively elaborate glucose isomerase under acid conditions typically unfavorable for growth of conventional glucose isomerase producing Streptomyces. Sterilization of the culture and production media may be avoided by selectively propagating newly discovered Streptomyces acidodurans under acidic conditions which will effectively eliminate contaminating microorganisms. The Streptomyces acidodurans herein also have the ability to undergo cultivation and elaborate glucose isomerase over a relatively broad pH range. Constitutive streptomycetes strains have also been isolated. Glucose isomerases derived from these Streptomyces strains are particularly effective for isomerizing glucose syrups to fructose-containing syrups.

Abstract: Disinfecting of immobilized enzymes is carried out by contacting the immobilized enzymes with a dilute aqueous solution of at least one substituted diethylenetriamine at a concentration and for a period of time which is sufficient to substantially kill the contaminating microorganisms without significant deleterious effects on the immobilized enzymes. The substituted diethylenetriamine is preferably dioctyldiethylenetriamine or a mixture of dioctyldiethylenetriamine and trioctyldiethylenetriamine.

Abstract: Whole microbial cells containing enzymes not sensitive to glutaraldehyde such as sucrose mutase or glucose isomerase are immobilized by forming a reaction product of the cells in an aqueous medium with glutaraldehyde, reacting the reaction product with polyethylenimine to flocculate the reaction product, and recovering the flocculated reaction product from the aqueous medium. Reacting the cells with glutaraldehyde prior to reacting with polyethylenimine results in flocculated cells that can be more easily separated from the aqueous medium.

Abstract: A novel strain of Bacillus licheniformis, ATCC 31667, capable of producing glucose isomerase in the absence of xylose is disclosed. Also disclosed is a novel method for screening Bacillus mutants for this property.

Abstract: There is provided a process for producing glucose-isomerase in high yield by culturing under aerobic conditions, the strain Streptomyces griseoflavus (NCIB 11542).

Abstract: A novel glucose isomerase enzyme useful for the conversion of glucose to fructose can be prepared by growing under aerobic conditions a culture of Bacillus licheniformis ATCC 31604 in a medium containing appropriate nutrients and then recovering the enyzme therefrom.

Abstract: A method of obtaining glucose isomerase which comprises cultivating the enzyme-producing strain Streptomyces sp. N.765, registration No. 143 (Bulgarian State Institute for Drug Control, Sofia, Bulgaria,) for 36 to 72 hours at a temperature of 24.degree. to 36.degree. C. at an initial pH of 6.5 to 9.0 in a culture medium containing 1.0 to 2.0% xylose 1.5 to 4.0% of dry weight of maize extract and 0.23 to 1.0% weight sodium acetate.

Abstract: Enzyme purification is carried out by contacting an impure liquid enzyme preparation containing enzyme and soluble impurities with an ion exchange material in a column to adsorb both the enzyme and impurities by the ion exchange material, adding an additional amount of the impure liquid enzyme preparation whereby the soluble impurities therein are preferentially adsorbed by the ion exchange material and the adsorbed enzyme is displaced from the ion exchange material to produce a purified liquid enzyme preparation containing higher enzyme activity than before purification. The purified enzyme is more highly adsorbed by ion exchange material when immobilizing the enzyme.

Abstract: An immobilized glucose isomerase is prepared by treating an aqueous suspension of cells of a glucose isomerase producing microorganism with a non-ionic surfactant that solubilizes glucose isomerase contained by the cells without solubilizing polysaccharides in the cells, separating the cells from the aqueous suspension, and adsorbing glucose isomerase from the resultant solution onto an ion exchange resin. By this process, contaminating polysaccharides are eliminated that inhibit adsorption of glucose isomerase on the ion exchange resin.

Abstract: A method for producing glucose isomerase comprising the steps of cultivating the enzyme producing strain Streptomyces sp. 1339 registration N. 144 Bulgarian State Institute for Drug Control, for 36 to 72 hours in a culture medium with xylose as an indicator, the temperature being kept at 24.degree. to 36.degree. C., the initial pH of cultivation being from 6.5 to 9.0 and effective isomerization at a temperature of 50.degree. to 80.degree. C., and a pH of 6.0 to 9.0 in the presence of 1.times.10.sup.-4 M CoCl.sub.2.6H.sub.2 O 1.times.10.sup.-2 M MgSO.sub.4.7H.sub.2 O in 0.1 to 3 M of the substrate.

Abstract: Fructose productivity and isomerase activity in immobilized beds or column operations employing isomerases obtained from Bacillus organisms are significantly improved by isomerizing a high solids feed syrup at pH 7.0-7.5 and 55.degree. C. to 60.degree. C. Without adding cobalt to the feed streams, continuous column operation in excess of 4,000 hours and yielding greater than 3,500 pounds of a 42% fructose syrup for each pound of isomerase can be achieved.

Abstract: The production of a heat stable glucose isomerase from a microorganism belonging to the genus Ampullariella and a method for using the isomerase to convert glucose to fructose.

Abstract: A method is disclosed for the production of fructose and syrups containing fructose and glucose, comprising the step of contacting a solution of glucose with a micro-organism of the genus Streptomyces sp. and more particularly of the strains NRRL 11.120 and NRRL 11.121, as designated by the Northern Regional Research Center, U.S. Department of Agriculture, Peoria, Ill.

Abstract: When immobilizing enzymes such as glucose isomerase by adsorption on a carrier such as an ion exchange resin, the enzyme prior to adsorption on the carrier is separated from a high molecular weight, non-dialyzable polysaccharide which inhibits adsorption of the enzyme to the carrier.

Abstract: A process for purifying an enzyme such as glucose isomerase which comprises precipitating nucleic acids from a cell-free, heat-treated enzyme solution in a suitable buffer and chromatographing the supernatant on a cellulosic medium. Subsequent chromatography on a hydrophilic, molecular-sieve medium affords enzyme of about 90% purity.

Abstract: In a process for extracting glucose isomerase from microorganism cells containing glucose isomerase by the so called high pressure release and impact method in which the suspension of the microorganism cells is homogenized for the extraction by releasing the pressure applied on said suspension instantaneously and giving the suspension an impact at very high velocity, the concentration of the suspension containing microorganism cells and the pressure applied on the said suspension are selected within specific ranges, namely from 0.5 to 10 wt. % (on a dry substance base) and from 400 to 700 Kg/cm.sup.2, respectively.

Abstract: A composition comprising intracellular or extracellular glucose isomerase may be purified by a method comprising heat treatment at a temperature from about 40.degree. C. to about 80.degree. C. The resultant enzyme solution, when utilized to prepare an immobilized enzyme system, is operationally equivalent to glucose isomerase purified by the traditional physico-chemical methods.