Abstract: Modified heparinases having altered binding specificity and activity are provided. Isolated nucleic acids encoding the same as as vectors and host cells are provided. Methods for using the modified heparinases are also provided.

Abstract: The cloning of the heparinase gene from Flavobacterium Heparinum using the polymerase chain reaction is described. The Open Reading Frame (ORF) corresponded to 1152 base pairs encoding a precursor protein of MW 43,800 daltons. The amino acid sequence reveals a 20-residue leader peptide. The gene was expressed in two expression systems in E. coli.

Abstract: A computer-implemented process determines candidate three-dimensional shapes of ring-shaped molecules through an exhaustive search of combinations of torsion angles of the bonds of the ring, given known bond lengths and bond angles for the bonds between atoms of the rings, a range of torsion angles and a resolution. Candidate combinations of the torsion angles are selected according to the range and resolution and coordinates of the atoms of the ring are determined for each candidate combination. It is then determined whether each candidate combination defines a physically possible ring. Indications of such candidate combinations are recorded. Candidate combination may then be selected from those recorded so that coordinates of atoms in candidate repetitive structures may be determined.

Abstract: A single, reproducible scheme to simultaneously purify all three of the heparin lyases from F. heparinum to apparent homogeneity is disclosed herein. The kinetic properties of the heparin lyases have been determined as well as the conditions to optimize their activity and stability. Monoclonal antibodies to the three heparinases are also described and are useful for detection, isolation and characterization of the heparinases.

Abstract: Pharmaceutical compositions for delivering an effective dose to a desired site of a heparinase. These compositions are based on the discovery that heparinase alone can inhibit angiogenesis. The effective dosage is dependent not only on the heparinase, but also on the method and means of delivery, which can be localized or systemic. For example, in some applications, as in the treatment of psoriasis or diabetic retinopathy, the inhibitor is delivered in a topical ophthalmic carrier. In other applications, as in the treatment of solid tumors, the inhibitor is delivered by means of a biodegradable, polymeric implant.

Abstract: A purified heparinase I, II and III free of lyase activity and each having a molecular weight of 42,800 84,100, 70,800, respectively, are produced by culturing Flavobacterium heparinum. The kinetic properties of the heparinases have been determined as well as the conditions to optimize their activity and stability.

Abstract: The present invention is an improved process for purification of active heparinase and heparinase like enzymes from Gram negative organisms, in particular, Flavobacterium heparinum. The primary advantage of the process is the fact that it allows large scale processing and high yield of heparinase. The heparinase is released from the periplasmic space of the organism by osmotic shock treatment, first into an osmotically stabilized medium, secondly into a non-stabilized medium having a pH of approximately pH 6.0 and 8.6 with subsequent release into a second non-stabilized medium containing approximately 0.15 M sodium chloride, followed by fractionation by cation exchange chromatography, and, optionally, electropheresis or gel filtration chromatography. Two proteins having heparinase activity have been isolated, one having a molecular weight of approximately 42,000 Daltons and the other having a molecular weight of 65,000 to 75,000 Daltons.

Abstract: In the membrane filtration of a liquid cell culture medium, superior flux rates and product recovery are obtained when a first charged particulate material and optionally a second charged particulate material bearing a charge opposite that of the first material are sequentially added to the medium prior to filtration.

Abstract: Apparatus for measuring L-glutamine in a liquid sample includes a membrane on which are immobilized the enzymes glutaminase and glutamate oxidase, whereby any said L-glutamine in the sample is acted upon by the glutaminase to form glutamic acid, the glutamic acid being acted upon by the glutamate oxidase to form an enzymatic reaction product, the membrane being operatively associated with a sensor capable of sensing the enzymatic reaction product or a compound or element consumed or liberated in the formation or degradation thereof.

Abstract: A process for forming a purified solute from an aqueous solution is provided whereby a mixture of an extractant, a hydrate former and the aqueous solution is first formed. The hydrate former forms a hydrate with water while the solute does not form a hydrate. The extractant takes up the solute from the aqueous solution. The mixture is subjected to a temperature and pressure sufficient to form the solid hydrate of the hydrate former, an aqueous solution of the solute and a portion comprising the extractant containing the solute. The solute is recovered from the portion comprising the extractant and the solute.

Abstract: Heparinase is produced by growing the bacterium, Flavobacterium heparinum in an improved defined medium consisting of a carbon source, two or more amino acids and several salts in the absence of protein. The carbon source concentration is specifically kept below a certain level to promote improved heparinase production. The sulfate source concentration is also specifically kept below a certain level to promote improved heparinase synthesis. Heparinase can be produced in this medium with or without the addition of an inducer compound.

Abstract: There is disclosed a heparin product obtained by degradation of heparin with heparinase from Flavobacterium heparinum (ATCC 13125) or mutants thereof having activity to reduce the coagulation activity of factor X while not effecting the coagulation activity of thrombin.

Abstract: Maltase is produced by growing a mutant of the yeast strain Saccharomyces italicus capable of growing in a growth medium utilizing sucrose as the carbon source. Maltase production is optimized by controlling time of fermentation, growth medium composition and sonication time during maltase recovery.

Abstract: Blood containing heparin and treated extracorporeally is contacted with immobilized heparinase prior to being reintroduced into the patient.

Abstract: Heparinase is produced by growing the bacteria, Flavobacterium heparinum, in a defined medium consisting of a carbon source, two or more amino acids and mineral salts in the absence of protein. Heparinase is recovered by batch chromatography of the cell extract from hydroxylapatite by elution with sodium chloride and sodium phosphate buffer washes.

Abstract: Maltase is produced by growing a mutant of the yeast strain Saccharomyces italicus capable of growing in a growth medium utilizing sucrose as the carbon source. Mutants of Saccharomyces italicus capable of utilizing sucrose as a carbon source also are provided.

Abstract: A purified heparinase I, II and III free of lyase activity and each having a molecular weight of 42,800 84,100, 70,800, respectively, are produced by culturing Flavobacterium heparinum. The kinetic properties of the heparinases have been determined as well as the conditions to optimize their activity and stability.

Abstract: A single, reproducible scheme to simultaneously purify all three of the heparin lyases from F. heparinum to apparent homogeneity is disclosed herein. The kinetic properties of the heparin lyases have been determined as well as the conditions to optimize their activity and stability. Mono-clonal antibodies to the three heparinases are also described and are useful for detection, isolation and characterization of the heparinases.