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Enzymatic phosphorylation of streptomycin by extracts of streptomycin-producing strains of Streptomyces.

Extracts of post-exponential phase mycelia of Streptomyces bikiniensis ATCC 11062, and other streptomycin-producers, catalyze phosphorylation of streptomycin and dihydrostreptomycin with adenosine-5'-triphosphate. The phosphate is esterified with an -OH group of the streptidine moiety. It is suggested that O-phosphoryl-streptomycin might serve as an intracellular precursor of extracellular streptomycin or as a detoxification product of streptomycin or that it might serve an unknown physiological function in the producing organism.

Adenosine Triphosphate↗

Effects of streptomycin deprivation on enzyme synthesis in streptomycin-dependent Escherichia coli.

During growth of streptomycin-dependent strains of Escherichia coli in the absence of streptomycin (deprived growth), both constitutive and inducible synthesis of beta-galactosidase were preferentially inhibited. A similar preferential inhibition of constitutive and derepressed synthesis of alkaline phosphatase was observed. Catabolite repression accounted for part, but not all, of the inhibition of the inducible beta-galactosidase synthesis. Serological experiments indicated that that part of the inhibition specificially associated with streptomycin deprivation was not a result of the production of altered beta-galactosidase. It is suggested that during deprived growth the ribosomes of streptomycin-dependent bacteria become impaired in their ability to translate certain messages.

Alkaline Phosphatase↗

Nature of streptomycin resistance in enterobacteria from faeces of streptomycin-treated patients in Egypt.

Streptomycin-resistant enterobacteria were isolated from the faeces of 31 out of 35 patients under streptomycin therapy, in the University Hospital of Alexandria. The isolated bacteria were 26 Escherichia coli, 3 Klebsiella pneumoniae and 2 Enterobacter cloacae. Their resistance patterns have shown that 80% had multiple antibiotic resistance. The MIC of streptomycin for all strains was higher than 312 microgram/ml. Nearly 90% of these strains carried resistance plasmids. However, 3 strains possessed no plasmids and were assumed to be mutants. 60% of the plasmid-carrying strains have been able to transfer the resistance by conjugation. The rest have shown small plasmids, some of them were characterized by streptomycin and sulphonamide resistance and a molecular weight of 4.1 mega-daltons.

Drug Resistance, Microbial↗

[Streptomycin-3"-phosphotransferases from streptomycin-resistant cells of Escherichia coli strains].

Streptomycin-3"-phosphotransferases were isolated and purified from E. coli cells containing plasmids 836, pBS52 or R6K, which determine the microorganisms resistance towards streptomycin and dihydrostreptomycin. Phosphorylation of the 3"-hydroxylic group of dihydrostreptomycin was demonstrated by [13C]-NMR spectrometry. It was shown that streptomycin-3"-phosphotransferase, whose synthesis is determined by plasmid 836 (as well as by plasmid R6K), differs from the analogous enzyme, whose synthesis is operated by plasmid pBS52 in some properties, e. g. dependence of the initial reaction rate on concentrations of antibiotics and ATP, pH-optimum, sensitivity to the buffer ionic strength, stability, etc. Besides, the antiserum against streptomycin-3'-phosphotransferase detected by plasmid pBS52 does not produce cross immunological reactions with the other enzyme.

Dihydrostreptomycin Sulfate↗

Ribosomal proteins from streptomycin-resistant and dependent mutants, and revertants from streptomycin-dependence to independence in Bacillus subtilis.

Streptomycin-resistant and dependent mutants were isolated from Bacillus subtilis ATCC 6633. Ribosomal proteins were analysed from six such mutants with chromatography on carboxymethyl cellulose or phosphocellulose columns. An altered specific 30s ribosomal protein, 30B, could be detected in all of these mutants. Streptomycin-independent revertants were isolated from a streptomycin-dependent strain. Some of them had an altered 30s ribosomal protein, 30A, and some others had an altered 30C protein. It was shown that from the data of partial N-terminal amino acid sequences together with amino acid compositions and mobilities on two-dimensional gel electrophoresis of these proteins that 30A, 30B, and 30C proteins were homologous with S5, S12 and S4 of E. coli 30s ribosomal proteins, respectively.

Amino Acid Sequence↗

Analysis of ribosomal proteins in streptomycin resistant and dependent mutants isolated from streptomycin independent Escherichia coli strains.

Mutants resistant to (Str-R) or dependent on streptomycin (Str-D) were isolated from several streptomycin independent (Str-I) strains of Escherichia coli. From 90 of these mutants ribosomes were isolated and the ribosomal proteins analyzed by two-dimensional polyacrylamide gel electrophoresis. The results which are summarized in Tables 1-4 led to the following conclusions: a) The phenotype (Str-R) or Str-D) of the mutants isolated from the Str-I strains strongly depends on the parental strain. b) No other ribosomal proteins than S4, S5 and S12 seem to be altered by mutations leading to dependence on, independence from or resistance to streptomycin. c) The S4 proteins of the analyzed mutants belong to three groups. The ratio between the groups depends more on the origin of the mutants than on their phenotype. d)Eight new types of altered S4 proteins were detected. It is very likely that many, if not all, of the altered S4 proteins originated by frame shift mutations. e) Some of the mutants differ from the wild type by alterations in three ribosomal proteins (S4, S5 and S12). The alteration in one protein, S4, apparently compensates for that in another protein, S5, in such a way that the original phenotype is expressed. These mutants are therefore an excellent tool for studies at the molecular level on the interaction of ribosomal components within the particle.

Drug Resistance, Microbial↗

Biosynthesis of streptomycin. Enzymic oxidation of dihydrostreptomycin (6-phosphate) to streptomycin (6-phosphate) with a particulate fraction of Streptomyces griseus.

Resting cells and to a greater extent permeabilized cells of Streptomyces griseus can oxidize dihydrostreptomycin to streptomycin. The dihydrostreptomycin oxidoreductase activity was localized in the 100,000 X g particulate fraction. Sucrose density gradient centrifugation of the particulate suspension gave a band at a density of 1.09 which consisted mainly of membrane vesicles. This fraction had high dihydrostreptomycin oxidoreductase activity. S. griseus protoplasts also contain high oxidoreductase activity. These data are consistent with localization of the enzyme in the cell membrane. Dihydrostreptomycin and dihydrostreptomycin 6-phosphate can both serve as substrates for the oxidoreducatase, but the phosphate was the better substrate in the cell free system. Addition of cofactors was not required for the bound dihydrostreptomycin oxidoreductase. The electron acceptor for the oxidation is unknown. Oxidation of dihydrostreptomycin 6-phosphate to streptomycin 6-phosphate very probably represents the penultimate step in the biosynthesis of streptomycin.

Centrifugation, Density Gradient↗

Correlation between streptomycin resistance and symbiotic properties of Rhizobium. I. Conversion of spheroplastizing, effective R. trifolii strain B1 to avirulent rods with changed phage and antibiotic patterns after mutation to high level of streptomycin resistance.

Rhizobium trifolii strain B1, which is infective and fixes nitrogen during symbiosis with clover plants, shows a peculiar property to undergo morphological change during growth, i.e. rods are changing into spheroplast-like forms. Moreover, it failed to grow at 38 degrees. It was found that mutation to high level of streptomycin resistance (above 1000 microgram per/ml) caused loss of this property. Further studies showed that simultaneously with the changes in streptomycin resistance other features of this strain were also changed: infectivity for clover plants, sensitivity to high temperature, phages and antibiotics. Mutation to low level of streptomycin resistance did not change the above mentioned features of the strain B1.

Bacteriophages↗

Streptomycin perfusion of the labyrinth through the round window plus intravenous streptomycin.

Streptomycin perfusion of the labyrinth through the round window membrane in the middle ear is a simple, safe, and effective way to stop the dizzy spells, fullness, and tinnitus of Ménière's disease without making the hearing worse. My results with 24 patients in the last 8 months are presented. Although the long-term effect of this operation remains to be seen, my experience with streptomycin perfusion of the labyrinth through the lateral semicircular canal leads me to believe these good results will be maintained in the future. Although the caloric response is not usually eliminated completely in the operated ear, it is reduced enough that the patient no longer has dizzy spells. Because the vestibular receptors are not completely destroyed, and the afferent and efferent nerves coming to and from them are undamaged, the patient compensates after this operation more quickly than when the vestibular receptors are completely destroyed by intramuscular or middle ear aminoglycoside, or when the afferent and efferent nerves of the vestibular receptors are cut as in labyrinthectomy or vestibular neurectomy. Hyaluronan is used to carry the streptomycin into the inner ear because it is known to penetrate the round window easily, and because being hygroscopic, it may reduce the amount of endolymph by osmosis.(ABSTRACT TRUNCATED AT 250 WORDS)

Ear, Inner↗

Improved expression of streptomycin resistance in plants due to a deletion in the streptomycin phosphotransferase coding sequence.

Previous studies have shown that a chimeric streptomycin phosphotransferase (SPT) gene can function as a dominant marker for plant cell transformation. The SPT marker previously described by Jones and co-workers has a limited value since it conferred a useful level of resistance only to a fraction (10%) of Nicotiana plumbaginifolia transgenic lines. Expression of resistance was species specific: no such resistant transformants were found in N. tabacum. In this paper we describe an improved SPT construct that utilizes a mutant Tn5 SPT gene. The mutant gene, SPT*, encodes a protein with a two amino acid deletion close to its COOH-terminus. In N. tabacum cell culture the efficiency of transformation with the improved streptomycin resistance marker was comparable to kanamycin resistance. When the chimeric SPT* gene was introduced linked to a kanamycin resistance gene, streptomycin resistance was expressed in most of the transgenic N. tabacum lines.

Chimera↗

Derivation and properties of Proteus mirabilis systems for high frequency transduction of streptomycin--sulphonamide and streptomycin-sulphonamide--kanamycin resistances.

Properties of two transducing systems with phages capable of high frequency transduction (HFT) of streptomycin and sulphonamide resistance markers of the V group plasmid R905, and of these markers plus the kanamycin resistance marker derived from a previously described HFT phage 5006MHFTak, are described. Transducing particles of the former phage, named 5006MHFTsus, were detected using the replica-plate technique in an ultraviolet-induced lysate of Proteus mirabilis strain PM5006 transduced to streptomycin and sulphonamide resistance by phage 5006M grown on PM5006 carrying R905. Phage 5006MHFTsusk was also detected by the replica-plate technique in ultraviolet-induced lysates of of phage 5006MHFTsus transductants retransduced to ampicillin and kanamycin resistance by phage 5006MHFTak. Both phages were serologically identical to the parent phage 5006M. Ultraviolet-induced lysates transduced their markers to PM5006 at frequencies of about 5 X 10(-2)/plaque-forming unit adsorbed for both the phages. With phage 5006MHFTsusk, this frequency was increased about 10-fold by simultaneous infection of recipients with homologous non-transducing phage, while phage 5006MHFTsus transductions only underwent a twofold increase. Transductants took about 60 min to express complete resistance to 50 mug streptomycin/ml, and resistance to 1600 mug sulphadiazine/ml was complete within 120 min after phage adsorption. Phage 5006MHFTsusk was slightly more resistant to ultraviolet inaction of its transducing potential and reasons are given for the belief that transductants of both phages are heterogenote-like. Both phage lysates were also capable of generalized transduction and, like previously described HFT phages, lysates transduced the leucine marker at increased frequencies. Using previously described extra- and intra-species phages hosts, it was found that the phages could transduce in single infection and were defective in the lysogenic conversion function as well as in a maturation step. Possible modes of formation of the HFT particles are discussed9

Ampicillin↗

Two-dimensional gel electrophoresis of ribosomal proteins from streptomycin-sensitive and streptomycin-resistant mutants of Chlamydomonas reinhardi.

Ribosomal proteins from three mutant strains of Chlamydomonas reinhardi were analysed and compared by one-dimensional and two-dimensional gel electrophoresis. One mutant was streptomycin-sensitive the other two were streptomycin-resistant, one with a Mendelian the other with a non-Mendelian pattern of inheritance. In the 30-S subunits of chloroplast ribosomes approximately 25 proteins are found and in the 50-S subunits 34 proteins. The 40-S subunits of cytoplasmic ribosomes contain about 31 proteins and the 60-S subunits 44 proteins. The molecular weights of most proteins in all subunits are in the range of 10 000 to 35 000. However, the 60-S subunits contain in addition a protein of molecular weight 50 000 and the 30-S subunits show 6-7 bands of molecular weights from 50 000 to 83 000. The proteins of the cytoplasmic 80-S ribosomes or of their subunits from all three mutants are electrophoretically identical. The proteins of the 70-S organellar ribosomes and both of their subunits show distinct differences between the three strains. Our results indicate that organellar ribosomal proteins are in part controlled by nuclear DNA and in part by organellar DNA.

Chlamydomonas↗

Streptomycin-dependent exhibition of cytokine-inducing activity in streptomycin-dependent Mycobacterium tuberculosis strain 18b.

Peritoneal exudate cells of mice were stimulated with a streptomycin-dependent Mycobacterium tuberculosis strain, 18b. Gamma interferon production by natural killer cells depending on interleukin-12 and interleukin-18 was induced only in the presence of a high dose of streptomycin. This study suggested the requirement of active bacterial metabolism for this host response.

Animals↗

Treatment of streptomycin-susceptible and streptomycin-resistant enterococcal endocarditis.

Fifty-six patients with enterococcal endocarditis received 4 weeks of antimicrobial therapy with penicillin G and streptomycin (36 patients) or, if infections were streptomycin resistant, penicillin and gentamicin (20 patients). Compared with patients who had symptoms for less than 3 months, patients with symptoms for more than 3 months had a higher relapse rate (0% versus 44%; p less than 0.001) and mortality (2.5% versus 25%; p less than 0.001). Patients with mitral valve endocarditis had a significantly higher relapse rate (25%) than patients with aortic valve infections (0%) (p less than 0.01). Gentamicin-associated nephrotoxicity was more frequent (p less than 0.001) among patients treated with greater than 3 mg/kg d of gentamicin than among those treated with 3 mg or less (100% versus 20%). Relapse and mortality rates did not differ significantly between patients treated with low-dose or high-dose gentamicin regimens. Patients who have had symptoms of enterococcal endocarditis for more than 3 months or patients with mitral valve infection should receive at least 6 weeks of antimicrobial therapy, but patients without these high-risk factors can be treated for 4 weeks.

Adult↗

A second streptomycin resistance gene from Streptomyces griseus codes for streptomycin-3"-phosphotransferase. Relationships between antibiotic and protein kinases.

Two genes, aphE and orf, coding for putative Mr 29,000 and Mr 31,000, proteins respectively, were identified in the nucleotide sequence of a 2.8 kbp DNA segment cloned from Streptomyces griseus N2-3-11. The aphE gene expressed streptomycin (SM) resistance and a SM phosphorylating enzyme in S. lividans strains. The two genes were found to be in opposite direction and seemed to share a common region of transcription termination. The aphE gene shows significant homology to the aph gene, encoding aminoglycoside 3'-phosphotransferase, APH(3'), from the neomycin-producing S. fradiae. The enzymatic specificity of the aphE gene product was identified to be SM 3"-phosphotransferase, APH(3"). The primary structure of the APH(3") protein is closely related to the members of the APH(3') family of enzymes. However, the APH(3") enzyme did not detectably phosphorylate neomycin or kanamycin. There is only low similarity of the protein to the APH(6) group of SM phosphotransferases. An evolutionary relationship between antibiotic and protein kinases is proposed.

Amino Acid Sequence↗

[Immediate type anaphylaxis of streptomycin allergy elicited by streptomycin polymers].

It has been known that streptomycin (SM) can cause some immediate type anaphylaxis even anaphylactic shock in clinical therapy. The characteristic of the substance that elicits the allergic reaction has not been reported. Using gel filtration and high performance gel permeation chromatography (HPGPC), we have found that some impurities of high molecular weights (HMW) were formed by heating acidic SM solution. The HMW impurities had a colour reaction with citric acid-acetic anhydride reagent and elicited passive cutaneous anaphylaxis (PCA) on guinea pigs sensitized with rabbit anti-SM-BSA serum and general anaphylaxis in guinea pigs immunized by SM-BSA. It is suggested that the impurities, SM polymers (poly-SM) related to some reactions on amino groups of SM, are the allergens of SM allergy.

Animals↗

[Effect of the prolonged use of streptomycin as well as streptomycin in combination with tubazid on the bioelectrical activity of the brain in experimental tuberculosis].

The effect of streptomycin used alone and in combination with tubazid on the brain electric activity was studied in chronic experiments on rabbits with tuberculosis. The electrocorticographical investigations showed that the antibacterial treatment lowered the spontaneous electric activity and reactive capacity of the cortex. It was evident from a narrow amplitude of the main rythm, coefficient and energy of synchronization on rythmic photostimulation and paroxysmal activity withing the theta range. With the account of the high therapeutic effect of the antibacterial treatment evident from minimum affection of the organs with tuberculosis, the above changes in the electrocorticogrammes should be attributed to the neurotropic effect of the drug and not to tuberculosis intoxication.

Alpha Rhythm↗