The possible occurrence of zinc in ribitol and sorbitol dehydrogenases from Mycobacterium sp. 279

The activity of polyhydric alcohol dehydrogenases in Mycobacterium sp. 279 was studied under limitation of zinc in the growth medium. It was found that the activity of ribitol and sorbitol dehydrogenases were markedly lowered and that of D-arabinitol dehydrogenase remained unchanged in the Zn2+-deficient cells. Other ions tested i.e., Co2+, Cu2+, Ni2+ and Mn2+ failed to substitute Zn2+ ions in their effect on the enzyme activities. The Zn2+-responsive enzymes were sensitive to the chelating agents (1,10-phenanthroline, 2,2'-dipyridyl), whereas D-arabinitol dehydrogenase was insensitive. The results indicate possible existence of a zinc component in the ribitol and sorbitol dehydrogenases from Mycobacterium sp. 279.     

17-Hydroxysteroid dehydrogenases of Mycobacterium sp. VKM Ac-1815D mutant strain

Whole cells and crude extract of Mycobacterium sp. VKM Ac-1815D mutant strain Et1 were shown to carry out 17beta-reduction, 17beta-dehydrogenation and 1(2)-reduction of 3-keto-C(19)-steroids. Two 17-hydroxy steroid dehydrogenases (17-OH SDH) were partially purified from the strain by ammonium sulfate fractionation, ion-exchange chromatography on DEAE-sephacel and gel-filtration on Bio-Gel A. The enzymes differed in chromatographic properties and specific activities. One enzyme--17-OH SDH (2) (tetramer, M(r) approximately 210,000) was found to be responsible for bi-directional reduction-oxidation of steroids at C 17, whereas the other one--17-OH SDH (1) (monomer, M(r) approximately 68,000) specifically catalysed 17beta-dehydrogenation of 17-hydroxysteroids (testosterone and 1(2)-dehydro testosterone).The 17beta-reduction of 1-ene-17-ketosteroids was accompanied by 1(2)-reduction. A role of 1-ene-reductase as a steroid-binding protein associated with 17-OH SDH (2) in Mycobacterium sp. is discussed.     

Isolation and partial characterization of a new mutant for sterol biotransformation in Mycobacterium sp.

Summary A new sterol biotransforming mutant was isolated from NRRL-B3683 Mycobacterium sp. after nitrosoguanidine mutagenesis. The mutant showed an enhanced ability to biotransform stigmasterol into 17-ketosteroids compared with the parental strain.     

Localization of 17beta-hydroxysteroid dehydrogenase in Mycobacterium sp. VKM Ac-1815D mutant strain

The localization of mycobacterial 17β-hydroxysteroid dehydrogenase (17β-OH SDH) was studied using cell fractionation and cytochemical investigation. Mycobacterium sp. Et1 mutant strain derived from Mycobacterium sp. VKM Ac-1815D and characterized by increased 17β-OH SDH activity was used as a model organism.

Subcellular distribution study showed both soluble and membrane-bound forms of mycobacterial 17β-hydroxysteroid dehydrogenase. The cytochemical method based on a copper ferrocyanide procedure followed by electron microscopic visualization was applied in order to investigate the intracellular localization of bacterial 17β-OH SDH in more detail. The enzyme was found to be located in the peripheral cytoplasmic zone adjoining the cytoplasmic membrane (CM). 17β-OH SDH was loosely membrane bound and easily released into the environment under the cell integrity failure.     

Mycobacterium rhodesiae sp. nov.

A number of rapid-growing scotochromogenic mycobacteria were isolated from the sputum specimen of Rhodesian patients with pulmonary disease and recognized as a new species. This was given the name Mycobacterium rhodesiae sp. nov. A comparison between M. rhodesiae, M. parafortuitum, M. aurum and M. vaccae was done, and distinguishing characters serving for differentiation between these 4 species of rapid-growing scotochromogenic mycobacteria were described.     

Mycobacterium confluentis sp. nov.

A new rapidly growing mycobacterium was isolated from human sputum. This organism grew at 22, 31, 37, and 41 degrees C and possessed catalase, acid phosphatase, acetamidase, urease, nicotinamidase, pyrazinamidase, and nitrate reductase activities. It did not produce nicotinic acid, hydrolyze Tween, or have benzamidase, isonicotinamidase, succinidamidase, and arylsulfatase activities. A mycolic acid analysis revealed a simple, unique pattern. The organism is susceptible to antituberculotic drugs. A comparative 16S rRNA sequence analysis placed this organism within the confines of the genus Mycobacterium, most closely related to the thermotolerant rapidly growing species. On the basis of the pattern of enzymatic activities and metabolic properties, as well as the unique 16S rRNA sequence, we propose that our single strain represents a new species, for which we propose the name Mycobacterium confluentis. The type strain is strain 1389/90; a culture of this strain has been deposited in the German Collection of Microorganisms and Cell Cultures as strain DSM 44017.     

Mycobacterium alvei sp. nov.

A new species of rapidly growing, nonphotochromogenic mycobacteria, Mycobacterium alvei, is described. The inclusion of this organism in the genus Mycobacterium is based on its acid fastness, its mycolate pattern, and its G + C content. A study of six strains showed that they form a homogeneous group with an internal phenotypic similarity value of 97 +/- 2.22%. DNA relatedness studies showed that the six M. alvei strains which we studied form a single DNA hybridization group which is less than 49% related to 14 other species of the genus Mycobacterium; the deltaTm values determined for the strains which exhibited higher levels of DNA homology were all greater than 7.9 degrees C. A lipid analysis showed that tuberculostearic acid was present. Docosanoic and tetracosanoic acid methyl esters were detected as mycolic acid cleavage products. All six isolates which we tested contained alpha-mycolic acids and relatively large amounts of a new kind of mycolic acid containing a methoxy group of omega-1 position, a characteristic that has not been described previously in mycobacteria. Strain CR-21 is the type strain; a culture of this strain has been deposited in the Collection Nationale de Cultures de Microorganismes de l'Institut Pasteur, Paris, France, as strain CIP 103464.     

Production of testosterone from phytosterol using a single-step microbial transformation by a mutant of Mycobacterium sp.

A testosterone (TS)-producing mutant, ST2, was derived from a phytosterol-assimilating and androst-4-ene-3,17-dione (AD)-producing bacterium, Mycobacterium sp. B-3805S, using nitrosoguanidine (NTG) mutagenesis. Production of TS from phytosterol using a single-step microbial transformation process by ST2 was investigated in a 5-l surface-aeration microprocessor-controlled fermentor loaded with a synthetic medium supplemented with 0.1% phytosterol, 2% glucose and 1% peptone at 30°C. An increase in dissolved oxygen at the initial stage of fermentation favored the side-chain degradation of phytosterol to AD. Later in the fermentation, a decrease in the dissolved oxygen to zero resulted in a decrease in pH to 6.0 as well as the reduction of AD to TS. Under optimal fermentation conditions, the maximum conversion ratio of phytosterol to TS was 31% after 120 h cultivation. It was concluded that the control of dissolved oxygen in the fermentation culture is the most important parameter for production of TS from phytosterol via AD. TS was isolated from the fermentation culture by addition of Amberlite XAD-7 resin and was further purified by flash chromatography on a silica gel column. After crystallization, TS was obtained as needle crystals with the correct melting point. Journal of Industrial Microbiology & Biotechnology (2002) 28, 280–283 DOI: 10.1038/sj/jim/7000243 Received 20 September 2001/ Accepted in revised form 16 January 2002     

A mutant of Anabaena sp. CA with oxygen-sensitive nitrogenase activity.

Studies on the O₂ protection mechanism for nitrogenase in a mutant (PM10) of $\text{Anabaena}$ sp. CA indicated that the ability to protect nitrogenase from O₂ was functionally impaired. Growth rates of PM10 were substantially improved when cells were cultured under microaerobic conditions. Nitrogenase activity was totally inhibited by exposure to O₂ for 30 min; partial restoration of activity was attained when cell suspensions were subsequently made microaerobic. Experiments in which induction of nitrogenase activity was followed indicated that the synthesis of the O₂ protection mechanism was temporally separated from synthesis of heterocysts and nitrogenase.     

Mineralization of phenanthrene by a Mycobacterium sp.

A Mycobacterium sp., designated strain BG1, able to utilize the polycyclic aromatic hydrocarbon phenanthrene as the sole carbon and energy source was isolated from estuarine sediment following enrichment with the hydrocarbon. Unlike other phenanthrene degraders, this bacterium degraded phenanthrene via 1-hydroxy-2-naphthoic acid without accumulating this or other aromatic intermediates, as shown by high-performance liquid chromatography. Degradation proceeded via meta cleavage of protocatechuic acid. Different nonionic surfactants (Tween compounds) solubilized the phenanthrene to different degrees and enhanced phenanthrene utilization. The order of enhancement, however, did not correlate perfectly with increased solubility, suggesting physiological as well as physicochemical effects of the surfactants. Plasmids of approximately 21, 58, and 77 megadaltons were detected in cells grown with phenanthrene but not in those which, after growth on nutrient media, lost the phenanthrene-degrading phenotype. Given that plasmid-mediated degradations of aromatic hydrocarbons generally occur via meta cleavages, it is of interest that the addition of pyruvate, a product of meta cleavage, supported rapid mineralization of phenanthrene in broth culture; succinate, a product of ortho cleavage, supported growth but completely repressed the utilization of phenanthrene. The involvement of plasmids may have given rise to the unusual degradation pattern that was observed.