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Vanyushkina A.A.,RAS Research Center Kurchatov Institute | Kamashev D.E.,RAS Research Center Kurchatov Institute | Altukhov I.A.,Russian Institute of Physico Chemical Medicine | Govorun V.M.,RAS Research Center Kurchatov Institute
Biochemistry (Moscow) | Year: 2012

In contrast to the abundance of systems-oriented approaches describing changes on the transcriptome or pro- teome level, relatively few studies have employed the metabolome. The goal of the presented research was to identify as many intracellular metabolites as possible in a Spiroplasma melliferum extract by flow injection time-of-flight mass spec- trometry. The Mollicutes class bacterium S. melliferum is a member of a unique category of bacteria that have in common the absence of a cell wall, a reduced genome, and simplified metabolic pathways. Metabolite identification was confirmed by fragmentation of previously detected ions by target mass spectrometry. The selected liquid chromatography approach, hydrophilic interaction chromatography with amino and silica columns, effectively separates highly polar cellular metabo- lites prior to their detection on a high accuracy mass spectrometer in positive and negative acquisition mode for each col- umn. Here we present reliable measurement of 76 metabolites, including components of sugar, amino acid, and nucleotide metabolism. We have identified about a third of the possible intracellular S. melliferum metabolites predicted by genome annotation. © Pleiades Publishing, Ltd., 2012. Source

Vanyushkina A.A.,Russian Institute of Physico Chemical Medicine | Fisunov G.Y.,Russian Institute of Physico Chemical Medicine | Gorbachev A.Y.,Russian Institute of Physico Chemical Medicine | Kamashev D.E.,RAS Research Center Kurchatov Institute | Govorun V.M.,Moscow Institute of Physics and Technology
PLoS ONE | Year: 2014

We present a systematic study of three bacterial species that belong to the class Mollicutes, the smallest and simplest bacteria, Spiroplasma melliferum, Mycoplasma gallisepticum, and Acholeplasma laidlawii . To understand the difference in the basic principles of metabolism regulation and adaptation to environmental conditions in the three species, we analyzed the metabolome of these bacteria. Metabolic pathways were reconstructed using the proteogenomic annotation data provided by our lab. The results of metabolome, proteome and genome profiling suggest a fundamental difference in the adaptation of the three closely related Mollicute species to stress conditions. As the transaldolase is not annotated in Mollicutes, we propose variants of the pentose phosphate pathway catalyzed by annotated enzymes for three species. For metabolite detection we employed high performance liquid chromatography coupled with mass spectrometry. We used liquid chromatography method - hydrophilic interaction chromatography with silica column - as it effectively separates highly polar cellular metabolites prior to their detection by mass spectrometer. © 2014 Vanyushkina et al. Source

Alexeev D.,Moscow Institute of Physics and Technology | Kostrjukova E.,Russian Institute of Physico Chemical Medicine | Aliper A.,Russian Institute of Physico Chemical Medicine | Popenko A.,Russian Institute of Physico Chemical Medicine | And 15 more authors.
Journal of Proteome Research | Year: 2012

To date, no genome of any of the species from the genus Spiroplasma has been completely sequenced. Long repetitive sequences similar to mobile units present a major obstacle for current genome sequencing technologies. Here, we report the assembly of the Spiroplasma melliferum KC3 genome into 4 contigs, followed by proteogenomic annotation and metabolic reconstruction based on the discovery of 521 expressed proteins and comprehensive metabolomic profiling. A systems approach allowed us to elucidate putative pathogenicity mechanisms and to discover major virulence factors, such as Chitinase utilization enzymes and toxins never before reported for insect pathogenic spiroplasmas. © 2011 American Chemical Society. Source

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