Synthesis of melanin by a moderately thermophilic methanotroph Methylocaldum szegediense depends on cultivation temperature

Mutant variants of mini-intein PRP8 from Penicillium chrysogenum (Int4b) with improved control of C-terminal processing were characterized. The presented variants can serve as a basis for self-removed polypeptide tags capable of carrying an affine label and allowing to optimize the process of obtaining target proteins and peptides in E. coli cells. They allow to synthesize target molecules in the composition of soluble and insoluble hybrid proteins (fusions), provide their afnne purification, autocatalytic processing and obtaining mature target products. The presented variants have a number of features in comparison with the known prototypes. In particular the mutant mini-intein Int4bPRO, containing the L93P mutation, has temperature-dependent properties. At cultivation temperature below 30 °C it allows the production of target molecules as part of soluble fusions, but after increasing of cultivation temperature to 37 °C it directs the most of synthesized fusions into insoluble intracellular aggregates. The transition of Int4bPRO into insoluble form is accompanied by complete inactivation of C-terminal processing. Further application of standard protein denaturation-renaturation procedures enable efficiently reactivate Int4bPRO and to carry out processing of its fusions in vitro. Two other variants, Int4b56 and Int4b36, containing a point mutation T62N or combination of mutations D144N and L146T respectively, have a reduced rate of C-terminal processing. Their use in E. coli cells allows to optimize the biosynthesis of biologically active target proteins and peptides in the composition of soluble fusions, suitable for afnne purification and subsequent intein-dependent processing without the use of protein denaturation-renaturation procedures. intein, fusion, processing, processing rate, gelonin The work was supported within the framework of the State Assignment no. 595-00003-19 PR.

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Ferrous iron oxidation in moderately thermophilic acidophile Sulfobacillus sibiricus N1T

Canadian Journal of Microbiology ◽

10.1139/w10-063 ◽

2010 ◽

Vol 56 (10) ◽

pp. 803-808 ◽

Cited By ~ 10

Author(s):

Tatiana Y. Dinarieva ◽

Anna E. Zhuravleva ◽

Oksana A. Pavlenko ◽

Iraida A. Tsaplina ◽

Alexander I. Netrusov

Keyword(s):

Ferrous Iron ◽

High Performance ◽

Iron Oxidation ◽

Early Exponential Phase ◽

Moderately Thermophilic ◽

Terminal Oxidases ◽

Cytochrome Bo ◽

Ferrous Iron Oxidation ◽

Transport Chain ◽

Membrane Bound

The iron-oxidizing system of a moderately thermophilic, extremely acidophilic, gram-positive mixotroph, Sulfobacillus sibiricus N1T, was studied by spectroscopic, high-performance liquid chromatography and inhibitory analyses. Hemes B, A, and O were detected in membranes of S. sibiricus N1T. It is proposed that the electron transport chain from Fe2+ to O2 is terminated by 2 physiological oxidases: aa3-type cytochrome, which dominates in the early-exponential phase of growth, and bo3-type cytochrome, whose role in iron oxidation becomes more prominent upon growth of the culture. Both oxidases were sensitive to cyanide and azide. Cytochrome aa3 was more sensitive to cyanide and azide, with Ki values of 4.1 and 2.5 µmol·L–1, respectively, compared with Ki values for cytochrome bo3, which were 9.5 µmol·L–1 for cyanide and 7.0 µmol·L–1 for azide. This is the first evidence for the participation of a bo3-type oxidase in ferrous iron oxidation. The respiratory chain of the mixotroph contains, in addition to the 2 terminal oxidases, a membrane-bound cytochrome b573.

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Decomposition of Rice Chaff Using a Cocultivation System of Thermobifida fusca and Ureibacillus thermosphaericus

Proceedings ◽

10.3390/proceedings2020066031 ◽

2021 ◽

Vol 66 (1) ◽

pp. 31

Author(s):

Sachiko Nakamura ◽

Norio Kurosawa

Keyword(s):

Lignin Peroxidase ◽

Culture Medium ◽

Reducing Sugars ◽

Thermophilic Bacteria ◽

Thermobifida Fusca ◽

Bacterial Strains ◽

Basal Salt Medium ◽

Moderately Thermophilic ◽

Fuels And Chemicals ◽

Ureibacillus Thermosphaericus

Lignocellulosic biomass comprises cellulose, hemicellulose, and lignin and is a potential source of fuels and chemicals. Although this complex biomass is persistent, it can be cooperatively decomposed by a microbial consortium in nature. In this study, a coculture of the moderately thermophilic bacteria Thermobifida fusca and Ureibacillus thermosphaericus was used for biodegradation of rice chaff. The bacterial strains were incubated in modified Brock’s basal salt medium (pH 8.0) supplemented with yeast extract and rice chaff at 50 °C for 7 days. The concentration of reducing sugars and the enzymatic activities of laccase, lignin peroxidase, cellulase, and xylanase in the supernatant of the culture medium were measured every day. The concentrations of reducing sugars in solo cultures of T. fusca and U. thermosphaericus and a mixed culture of the two strains after 7 days of incubation were 0.047, 0.040, and 0.195 mg/mL, respectively, indicating that the decomposition of rice chaff was enhanced in the coculture. Based on the results, it is thought that the lignin surrounding the cellulose was decomposed by laccase and lignin peroxidase secreted from U. thermosphaericus, resulting in cellulose and hemicellulose in the rice chaff being easily decomposed by enzymes from T. fusca.

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Membrane Lipid Composition of the Moderately Thermophilic Ammonia-Oxidizing Archaeon “Candidatus Nitrosotenuis uzonensis” at Different Growth Temperatures

Applied and Environmental Microbiology ◽

10.1128/aem.01332-19 ◽

2019 ◽

Vol 85 (20) ◽

Cited By ~ 4

Author(s):

Nicole J. Bale ◽

Marton Palatinszky ◽

W. Irene C. Rijpstra ◽

Craig W. Herbold ◽

Michael Wagner ◽

...

Keyword(s):

Lipid Composition ◽

Strong Influence ◽

Thermal Spring ◽

Membrane Lipid ◽

Effect Of Temperature ◽

Environmental Niche ◽

Membrane Lipid Composition ◽

Moderately Thermophilic ◽

Wide Range ◽

Membrane Spanning

ABSTRACT “Candidatus Nitrosotenuis uzonensis” is the only cultured moderately thermophilic member of the thaumarchaeotal order Nitrosopumilales (NP) that contains many mesophilic marine strains. We examined its membrane lipid composition at different growth temperatures (37°C, 46°C, and 50°C). Its lipids were all membrane-spanning glycerol dialkyl glycerol tetraethers (GDGTs), with 0 to 4 cyclopentane moieties. Crenarchaeol (cren), the characteristic thaumarchaeotal GDGT, and its isomer (crenʹ) were present in high abundance (30 to 70%). The GDGT polar headgroups were mono-, di-, and trihexoses and hexose/phosphohexose. The ratio of glycolipid to phospholipid GDGTs was highest in the cultures grown at 50°C. With increasing growth temperatures, the relative contributions of cren and crenʹ increased, while those of GDGT-0 to GDGT-4 (including isomers) decreased. TEX86 (tetraether index of tetraethers consisting of 86 carbons)-derived temperatures were much lower than the actual growth temperatures, further demonstrating that TEX86 does not accurately reflect the membrane lipid adaptation of thermophilic Thaumarchaeota. As the temperature increased, specific GDGTs changed relative to their isomers, possibly representing temperature adaption-induced changes in cyclopentane ring stereochemistry. Comparison of a wide range of thaumarchaeotal core lipid compositions revealed that the “Ca. Nitrosotenuis uzonensis” cultures clustered separately from other members of the NP order and the Nitrososphaerales (NS) order. While phylogeny generally seems to have a strong influence on GDGT distribution, our analysis of “Ca. Nitrosotenuis uzonensis” demonstrates that its terrestrial, higher-temperature niche has led to a lipid composition that clearly differentiates it from other NP members and that this difference is mostly driven by its high crenʹ content. IMPORTANCE For Thaumarchaeota, the ratio of their glycerol dialkyl glycerol tetraether (GDGT) lipids depends on growth temperature, a premise that forms the basis of the widely applied TEX86 paleotemperature proxy. A thorough understanding of which GDGTs are produced by which Thaumarchaeota and what the effect of temperature is on their GDGT composition is essential for constraining the TEX86 proxy. “Ca. Nitrosotenuis uzonensis” is a moderately thermophilic thaumarchaeote enriched from a thermal spring, setting it apart in its environmental niche from the other marine mesophilic members of its order. Indeed, we found that the GDGT composition of “Ca. Nitrosotenuis uzonensis” cultures was distinct from those of other members of its order and was more similar to those of other thermophilic, terrestrial Thaumarchaeota. This suggests that while phylogeny has a strong influence on GDGT distribution, the environmental niche that a thaumarchaeote inhabits also shapes its GDGT composition.

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Heterogeneity of population of microorganisms grown in presence of iron oxide maghemite nanoparticles

EPJ Web of Conferences ◽

10.1051/epjconf/201818510002 ◽

2018 ◽

Vol 185 ◽

pp. 10002 ◽

Cited By ~ 2

Author(s):

Tatyana P. Denisova ◽

Elena V. Simonova ◽

Lubov A. Kokorina ◽

Evgenia N. Maximova ◽

Oleg M. Samatov ◽

...

Keyword(s):

Iron Oxide ◽

Intercellular Communication ◽

Maghemite Nanoparticles ◽

Cultivation Temperature ◽

Large White ◽

Laser Target ◽

Water Based

In this work γ-Fe2O3 MNPs were obtained by laser target evaporation and water based suspensions were prepared. Maximum permissive dose of iron in water (MPD) is 0.3 mg/L. It was found that 100 MPD dose of iron induces formation of non-typical colonies after 72 or 96 hours exposition: against a background of small black colonies large white colonies appeared due to a disruption in tyrosine synthesis. Multiple re-cloning of the white colonies grown with MNPs showed that they retained their properties both under standard conditions (temperature of 24 °C) and at the temperatures up to 37o C. E.nigrum grown with MNPs demonstrated very scant extension of small colonies at the cultivation temperature of 24o C, their growth was completely blocked at 37°C. Significant changes in the structure of the population were noted. First of all, large cells with pronounced aggregation were observed among the black colonies. These aggregates consisted of large cells connected to each other by matrix. In the white colonies the appearance of very long threadlike cells connecting different groups of the cells establishing an intercellular communication was evident. Fe2O3 MNPs induce an increase in the heterogeneity of the population, expressed as a change in morpho-physiological states.

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Identification of Poly(cis-1,4-Isoprene) Degradation Intermediates during Growth of Moderately Thermophilic Actinomycetes on Rubber and Cloning of a Functional lcp Homologue from Nocardia farcinica Strain E1

Applied and Environmental Microbiology ◽

10.1128/aem.72.5.3375-3382.2006 ◽

2006 ◽

Vol 72 (5) ◽

pp. 3375-3382 ◽

Cited By ~ 58

Author(s):

Ebaid M. A. Ibrahim ◽

Matthias Arenskï¿½tter ◽

Heinrich Luftmann ◽

Alexander Steinbï¿½chel

Keyword(s):

Thermophilic Bacteria ◽

Natural Rubber Latex ◽

Gel Permeation Chromatography ◽

Mycolic Acid ◽

Rrna Gene ◽

Ionization Mass ◽

Thermophilic Actinomycetes ◽

Moderately Thermophilic ◽

Nocardia Farcinica ◽

Different Strains

ABSTRACT The enrichment and isolation of thermophilic bacteria capable of rubber [poly(cis-1,4-isoprene)] degradation revealed eight different strains exhibiting both currently known strategies used by rubber-degrading mesophilic bacteria. Taxonomic characterization of these isolates by 16S rRNA gene sequence analysis demonstrated closest relationships to Actinomadura nitritigenes, Nocardia farcinica, and Thermomonospora curvata. While strains related to N. farcinica exhibited adhesive growth as described for mycolic acid-containing actinomycetes belonging to the genus Gordonia, strains related to A. nitritigenes and T. curvata formed translucent halos on natural rubber latex agar as described for several mycelium-forming actinomycetes. For all strains, optimum growth rates were observed at 50ï¿½C. The capability of rubber degradation was confirmed by mineralization experiments and by gel permeation chromatography (GPC). Intermediates resulting from early degradation steps were purified by preparative GPC, and their analysis by infrared spectroscopy revealed the occurrence of carbonyl carbon atoms. Staining with Schiff's reagent also revealed the presence of aldehyde groups in the intermediates. Bifunctional isoprenoid species terminated with a keto and aldehyde function were found by matrix-assisted laser desorption ionization-time-of-flight and electrospray ionization mass spectrometry analyses. Evidence was obtained that biodegradation of poly(cis-1,4-isoprene) is initiated by endocleavage, rather than by exocleavage. A gene (lcp) coding for a protein with high homology to Lcp (latex-clearing protein) from Streptomyces sp. strain K30 was identified in Nocardia farcinica E1. Streptomyces lividans TK23 expressing this Lcp homologue was able to cleave synthetic poly(cis-1,4-isoprene), confirming its involvement in initial polymer cleavage.

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