[A method for selecting Streptomyces spheroides mutants--producers of an exoprotease with fibrinolytic activity]. / Metod otbora mutantov Streptomyces spheroides--produtsentov ékzoproteazy s fibrinoliticheskoi aktivnost'iu.

The paper present a description of a fast method to obtain mutants of Streptomyces spheroides--supersynthetics of exoprotease. The primary culture spores being irradiated by UV-rays, the mutants are formed with presence in the cultural medium of own proteolytic enzyme added from without. The described method is based on the phenomenon of growth suppression in the primary culture spore by high concentrations of the own proteolytic enzyme. It permits selecting producers with high proteolytic activity of fibrinolytic proteases from the small number of mutants grown in these colonies.

[Streptomyces spheroides mutant deprepressed for exoprotease biosynthesis]. / Mutant Streptomyces spheroides, derepressirovannyi po biosintezu ékzoproteaz.

The effect of carbon, nitrogen and sulfur sources on the biosynthesis of exoproteases was studied with the parent Streptomyces spheroides strain 35 and its mutant M8-2. Addition of a carbon, nitrogen and sulfur source to the medium deficient in one of these elements did not repress the synthesis of exoproteases by the washed mycelium of the mutant as compared to the parent strain. Protein as a sole source of carbon, nitrogen and sulfur had no effect on the biosynthesis of exoproteases by the mutant. In contrast to the parent strain, the biosynthesis of exoproteases in the mutant was not controlled by metabolite repression.

[Streptomyces spheroides M8-2 strain--a producer of extracellular proteolytic enzymes possessing fibrinolytic and thrombolytic action]. / Shtamm Streptomyces spheroides M8-2--produtsent vnekletochnykh proteoliticheskikh fermentov, okazyvaiushchikh fibrinoliticheskoe i tromboliticheskoe deistvie.

Strain Streptomyces spheroides M8-2 was obtained as a result of ultraviolet light action on the S. spheroides spores strain 35. The mutant is characterized by the considerable increase (in 4-5 times) of the extracellular proteolytic enzymes which are able to dissolve fibrin and thrombs. The description of the morfologic, cultural and physiologic-biochemical properties of the culture is given. The content of GC-pairs in DNK of the mutant and parent strains has been determined.

[Natural variability of Streptomyces spheroides--a producer of extracellular proteolytic enzymes possessing fibrinolytic action]. / Estestvannaia izmenchivost' Streptomyces spheroides--produtsenta vnekletochnykh proteoliticheskikh fermentov, obladaiushchikh fibrinoliticheskim deistviem.

Streptomyces spheroides M 8-2 was obtained using UV irradiation of spores taken from the parent culture. It shows a high activity of exocellular proteases which are capable of fibrin hydrolysis and degradation of blood clots. On certain media, a population of S. spheroides M8-2 was shown to yield three variants differing in their morphologo-biochemical properties. During submerged cultivation, these variants produce different quantities of exocellular proteases (3 to 19 units per 1 mg of biomass) with the fibrinolytic activity. Variants I and III are most active; their proteolytic activity is 10-14 units per 1 mg of biomass, on the average, and can reach 18-19 units per 1 mg of biomass. Variants with a low activity (variant II) are accumulated when the actinomycets is kept as spores on a solid medium with corn extract and on a solid medium with fibrin. The high proteolytic activity of the strain can be preserved by selection on a diagnostic medium with fibrin taking account of the diameter of hydrolytic zones around the colonies and selecting solely the colonies of variants I and III.

[Comparative study of proteolytic enzyme preparations of the actinomycete Streptomyces spheroides and its mutant]. / Sravnitel'noe izuchenie preparatov proteoliticheskikh fermentov aktinomitseta Streptomyces spheroides i ego mutanta.

Preparations of proteolytic enzymes with high fibrinolytic and thrombolytic activities were isolated from the cultural broth filtrates of Streptomyces spheroides and its mutant by precipitation with ammonium sulfate, dialysis and freeze-drying. The fibrinolytic and thrombolytic activities of the enzymes produced by the mutant are 5.6 and 6 times higher, respectively, than those of the parent strain. Moreover, the enzymes of the mutant have an elevated stability in the alkaline pH region, the optimum of their activities is also shifted to the alkaline pH region, they are more resistant to the action of blood plasma inhibitors and more sensitive to the action of high temperatures. As was shown using isoelectric focusing in a sucrose density gradient, the preparations of both the parent and mutant strains contain several proteolytic enzymes (or their groups) with acid, neutral and alkaline isoelectric points.

[Selection of Streptomyces spheroides--a producer of extracellular proteases with fibrinolytic action]. / Selektsiia Streptomyces spheroides--produtsenta vnekletochnykh proteaz s fibrinoliticheskim deistviem.

The mutagenic action of UV on Streptomyces spheroides 35 spores produced mutants with an elevated biosynthesis of exocellular proteases having the fibrinolytic activity. During submerged cultivation, mutant M8-2 accumulated 4-5 times more fibrinolytic enzymes with a high thrombolytic activity in the cultural broth than the parent strain. The selection of mutants synthesizing proteases with a high fibrinolytic activity can be made more effective by using a medium which contains fibrin, a substrate specific for the enzymes, as a sole source of carbon and nitrogen. The diameters of zones of fibrin hydrolysis around the colonies on this medium are a reliable criterion in the selection of such mutants. The preparation of proteolytic enzymes isolated from the cultural broth of mutant M8-2 is 5.3 and 3.9 times more active in fibrin and casein hydrolysis, respectively, than the preparation of the parent strain. Moreover, it has a high thrombolytic activity.