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Biomedical subjects

J Novotná

Publications and source records attributed to J Novotná.

At least 19 recordsLinked to original sources

Characteristics of Streptomyces globisporus strain 0234A forming endospores in submerged cultures.

Thermosensitive submerged endospores formed by Streptomyces globisporus 0234 and its natural variant A resembled those of thermoresistant actinomycetes not only in their morphology ultrastructure, but also in the content of dipicolinic acid. The production of endospores containing this substance is unusual in Streptomyces while other features of the strain indicate relatedness to other streptomycetes. Chemotaxonomic analysis of variant A revealed the cell wall to be of chemotype I and fatty acid content typical of Streptomyces. Most characteristics of surface cultures of variant A coincided with those of the original strain 0234 and its endosporeless variant B. Both the strain 0234 and its variants A and B produced identical antibiotics and pesticidal compounds.

Anti-Bacterial Agents

Characterization of an asporogenous mutant of Streptomyces aureofaciens with increased resistance to several aminoglycoside antibiotics.

An asporogenous spontaneous mutant of Streptomyces aureofaciens named ASR1 was selected on streptomycin gradient plates. The mutant is very stable and differs in ultrastructure and morphology, it is prototrophic but it lost the ability to grow well on soybean extract medium and produces one-tenth tetracyclines of the parent. The ASR1 mutant has a 3-4-fold increased resistance to streptomycin and is cross-resistant to other aminoglycosides. Comparison of the protein profiles from both strains on SDS gels revealed a very low expression of a 29.5 kDa protein in the ASR1 mutant which is overexpressed in both vegetative cells and spores of the parental strain.

Aminoglycosides

[Possibilities of predicting serum levels of lithium and subsequent correction of its dosage].

The objective of the present work was to evaluate the practicability of several procedures used for prediction of the serum concentration of lithium in a steady state which will make it possible to adjust the dosage in the early period of treatment. The group comprised nine men where prophylactic lithium treatment was indicated on account of maniomelancholy. The authors assessed in each patient the theoretically calculated and actually assessed value of minimal lithium concentrations in a steady state. In all applied predictive methods the predicted values were always higher than the actually assessed lithium concentrations. The least suitable method was that based on individualization of the population average of the velocity constant of elimination. The predictive value of the procedure based on a non-individualized single-point estimate was higher. A statistically significant correlation of predicted and assessed values was provided by the method which, based on two collected samples of lithium serum concentrations, makes it possible to estimate the individual value of the cumulative factor.

Adult

[The relation of nucleoli to precancerous conditions of the cervix].

Oestrogens act at the nuclear level. To these steroids also a cancerogenic effect is ascribed. By staining for nuclear structures according to Smetana and Busch the authors proved morphological structures of the nucleolus in vaginal epithelia and in cells from cervical lesions. This applies to solid nucleoli -- which are active, annular nucleoli which are resting and nucleoli which have completed their activity -- micronucleoli. Hyperoestrogenic women have in their vaginal epithelia of the upper third of the vagina more active nucleoli (11.4%) than hypooestrogenic women (2.1%). Women suffering from cervical carcinoma during the menopause have more active nucleoli in the vaginal epithelia (30.8%), as compared with healthy hyperoestrogenic women of productive age. Patients during the menopause with grade III dysplasias have fewer active nucleoli in cervical epithelia than women with cervical cancer (25.0%), however, as compared with hyperoestrogenic women of productive age, much more. A large number of active nucleoli was proved by staining by Smetana's method also in cells taken from cervical lesions carcinoma type and grade III dysplasia in postmenopausal patients. Evaluation of morphological changes of the nucleoli suggests their activity, the activity of the whole nucleolus and is a sensitive indicator of the activity of the whole cell.

Female

A direct-injection reversed-phase liquid chromatographic micromethod for studying the kinetics of terminal reactions of tetracycline biosynthesis.

A new micromethod for measuring enzyme-catalyzed reactions was developed. The method involves a number of consecutive direct injections of aliquots of the reaction mixture onto a microbore column and permits the determination of the time dependence of the decrease of substrate or increase of product concentrations. The reactions proceed in a microvial placed in the autosampler, and as the starting volume can be as low as 10 microliters, the requirement for the amount of enzyme is very low. The autosampler backed by the liquid chromatographic software allows automation of the analyses including data processing and easy quantitation of the enzymatic reaction(s). The method was applied to a system of two consecutive terminal reactions of tetracycline biosynthesis in Streptomyces aureofaciens, catalyzed by anhydrotetracycline oxygenase and tetracycline dehydrogenase. The usage of a diode-array detector facilitated the quantification of the reactions as the product and substrate could be monitored at their optimal wavelengths.

Chromatography, High Pressure Liquid

[Experimental models of arthritis induced by various cartilage collagen type].

To laboratory rats of Wistar strain (Ipcv: Wist) native collagenous types of pig cartilage--II, IX and XI and the denatured type XI were administered. The first immunization dose, 0.5 mg collagen per experimental animal, was administered in incomplete Freund adjuvant and the second immunization dose (collagen in solution) followed after 7 days and was administered by the i.p. route. Inflammatory arthritis of the small interphalangeal, metatarsophalangeal and tarsal joints developed between the 11th and 17th day of the experiment in rats immunized with native collagen type II and XI--60% of the animals were affected. The histological findings corresponded to the course of arthritis, however, the articular destructive changes after administration of collagen type XI were more severe than after type II. The changes were evaluated by X-ray and by means of thermovision. The antibody levels against different types of collagen assessed by means of the ELISA test reached peak values between the 13th and 20th day of the experiment. In the group of rats immunized by collagen type IX and the denatured type XI arthritis did not develop. The antibody response in the group of rats immunized with type IX was similar as in the group immunized with types II and XI. Histological examination confirmed some changes in the joints of rats immunized with collagen type IX, while in the joints of rats immunized with denatured collagen type XI no changes were found.

Animals

[The effect of a gold complex on experimental arthritis induced by immunization with type II collagen].

Gold complexes are used in the treatment of rheumatoid arthritis for some 60 years by now. The authors used therefore a gold complex, sodium aurothiosulphate (ATSS) to influence the experimental model of arthritis induced by immunization with type II collagen in laboratory rats. To the first group of laboratory rats ATSS was administered concurrently with the first immunization dose, to the second group with the second immunization dose and to the third group in the course of arthritis. ATSS was administered to individual groups every week by the i. m. route, 20 mg/kg body weight. In all three groups a reduction of arthritic symptoms was observed, however, in group three to a much lesser extent than in groups one and two. The results of the experiment indicate clearly that ATSS was able to suppress the development of collagen induced arthritis, if administered not later than with the second immunization dose. As the formation of antibodies against type II collagen was not suppressed, it may be assumed that the activation of the complement system was blocked by the bond of the gold complex with the C1q component. It has been proved already previously that the interaction of C1q with gold complexes is very rapid.

Animals

[The incidence of antibodies against collagen in patients with juvenile chronic arthritis].

Using the ELISA method, the authors assessed titres of antibodies to collagen type I, II and III in serum of patients with juvenile chronic arthritis (JCA). The results were compared with antibody titres in patients with rheumatoid arthritis (RA). Elevated antibody titres to all collagen types were found, as compared with age-matched controls. The mean titres in patients with RA were higher than in the group with JCA. Investigation of a correlation between the collagen antibody titres and values of the sedimentation rate after one hour revealed a statistical relationship only in collagen type II in patients with RA.

Adolescent

[Comparison of 2 methods for determining antibodies to collagen: the ELISA test and passive hemagglutination].

For the determination of antibodies against collagen in different rheumatic diseases the authors elaborated two serological techniques. It was particularly the passive haemagglutination, which proved to be little sensitive and insufficiently reproducible. Therefore, for the determination of antibodies against collagen the authors introduced the ELISA method as one of the varieties of enzyme immunoanalysis, giving more precise results. Both methods were compared and it has become apparent that the ELISA method is more reliable and more suitable for the determination of antibodies against collagen.

Autoantibodies

Isolation of pure anhydrotetracycline oxygenase from Streptomyces aureofaciens.

Anhydrotetracycline oxygenase was purified to homogeneity from Streptomyces aureofaciens, a producer of tetracycline. The enzyme was purified 60-fold in a 40% yield by a two-step procedure using a combination of hydrophobic chromatography and ion-exchange h.p.l.c. Purified anhydrotetracycline oxygenase was homogeneous according to SDS/polyacrylamide-gel electrophoresis, isoelectric focusing, ion-exchange h.p.l.c. on a Mono Q HR 5/5 column and size-exclusion h.p.l.c. on a TSK G 3000 SW column. The enzyme consists of two subunits of Mr 57,500, as determined by SDS/polyacrylamide-gel electrophoresis.

Chromatography, High Pressure Liquid

Anticollagen antibodies in patients with juvenile chronic arthritis.

Using ELISA antibodies, titres to collagen type I, II and III in sera of patients with juvenile chronic arthritis (JCA) were assayed. Results were compared with titres of patients with rheumatoid arthritis (RA). Increased antibodies titres to all three collagen types were found in comparison with healthy individuals of the same age. Average titres in patients with RA were higher than those of the JCA group. When antibodies titres were correlated with erythrocyte sedimentation rate in 1 hour (ESR/1h) statistical significance was found only in RA group for collagen type II.

Adolescent

Glucose-6-phosphate dehydrogenase from a tetracycline producing strain of Streptomyces aureofaciens: some properties and regulatory aspects of the enzyme.

Glucose-6-phosphate dehydrogenase from Streptomyces aureofaciens exhibited activity with both NAD and NADP, the maximum reaction rate being 1.6 times higher for NAD-linked activity than for the NADP-linked one. The KM values for NAD-linked activity were 2.5 mM for glucose-6-phosphate and 0.27 mM for NAD, and for NADP-linked activity 0.8 mM for glucose-6-phosphate and 0.08 mM for NADP. NAD- and NADP-linked activities were inhibited by both NADH and NADPH. (2'-phospho-)adenosinediphospho-ribose inhibited only NAD-linked activity. The inhibition was competitive with respect to NAD and noncompetitive with respect to glucose-6-phosphate.

Cations, Divalent

Partial purification and characterization of anhydrotetracycline oxygenase of Streptomyces aureofaciens.

Anhydrotetracycline oxygenase was purified both by affinity chromatography and by hydrophobic interaction chromatography. Molecular weight of anhydrotetracycline oxygenase was determined to be 115,000 by Sephadex G-200 gel filtration. Using preparative isoelectric focusing the isoelectric point of the enzyme was estimated to be 5.3. The enzyme showed a sensitivity to thiol-specific inhibitors. During the hydrophobic interaction purification step, the activity dropped considerably. Reactivation occurred when a heat treated crude extract was added to the reaction mixture.

Chromatography, Affinity

Subcellular localization of enzymes in Streptomyces aureofaciens and its alteration by benzyl thiocyanate. I. Phosphatases and ATP-glucokinase.

Mycelia of a low- and a high-production strain of Streptomyces aureofaciens were converted into protoplasts and divided into five subcellular fractions in order to localize exopolyphosphatases (EC 3.6.1.11), triphosphatase (EC 3.6.1.25), inorganic diphosphatase (EC 3.6.1.1), apyrase (EC 3.6.1.5) and glucokinase (EC 2.7.1.2). The highest specific activity of enzymes hydrolyzing polyphosphates was found in cytoplasmic vesicles and membranes. Triphosphatase was detected in the periplasmic fraction. Periplasmic vesicles and cytoplasm exhibited a high activity of diphosphatase. Apyrase was found mainly in the fractions of membranes and cytoplasmic vesicles. Glucokinase was a cytoplasmic enzyme. The enzymes were released from membrane structures into cytoplasm or periplasmic space if benzyl thiocyanate (10 microM) was present in the growth medium.

Cations

Subcellular localization of enzymes in Streptomyces aureofaciens and its alteration by benzyl thiocyanate. II. Anhydrotetracycline oxygenase and glucose-6-phosphate dehydrogenase.

The localization of anhydrotetracycline oxygenase and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) was studied by determining the enzyme activities in subcellular fractions obtained by differential centrifugation of the mycelia of Streptomyces aureofaciens after lysozyme treatment. Glucose-6-phosphate dehydrogenase was a typical cytoplasmic enzyme both in the low- and high-production strain. Anhydrotetracycline oxygenase was found in the membrane fraction of the low-production strain. In the high-production strain, it was detected in several fractions, the highest activity being found in cytoplasm. The presence of 10 microM benzyl thiocyanate in the culture medium significantly changed the distribution of the latter enzyme in both strains. The redistribution of the enzymes is discussed with respect to tetracycline over-production.

Cell Fractionation