Фенольные соединения относятся к веществам вторичного (специализированного) метаболизма, состав и... more Фенольные соединения относятся к веществам вторичного (специализированного) метаболизма, состав и содержание которых наиболее изучены в высших растениях, в отличие от прокариот. В настоящей работе представлены данные по изучению накопления этих метаболитов в процессе роста цианобактерий Synechocystis sp. (Synechocystis sp. PCC 6803 GT-L, штамм IPPAS В-1400) и Desertifilum tharense (штамм IPPAS B-1220) из коллекции микроводорослей и цианобактерий IPPAS ИФР РАН, а также исследованию их состава. Для этого были использованы методы спектрофотометрического анализа, тонкослойной хроматографии, УФ-спектрометрии. Установлено более высокое накопление фенольных соединений у Synechocystis sp., которое почти вдвое превышало таковое у D. tharense. В период линейной фазы роста (3 сутки) оно было выше такового в стационарную фазу роста (10 сутки) у обеих культур. Выявлены отличия в составе фенольных соединений, присутствующих в этанольных экстрактах Synechocystis sp. и D. tharense (4 и 7 веществ соответственно). Установлено наличие в них коньюгатов п-оксибензойной и п-кумаровой кислот − метаболитов начальных этапов биогенеза фенольных соединений.
Вестник Российского университета дружбы народов. Серия: Агрономия и животноводство, 2010
This article dedicates the study concerning evolution of the hormonal system of plants, in partic... more This article dedicates the study concerning evolution of the hormonal system of plants, in particular, detection of components of the regulatory system of abscisic acid (ABA) in cyanobacteria, which are considered as the precursors of the chloroplasts. ABA-binding proteins with molecular masses of 23, 50, 60 and 67 kDa were isolated using affinity chromatography on Sepharose conjugated with ABA from the lysate of cyanobacteria Synechocystis sp. PCC 6803. Interaction of isolated proteins with idiotypic antibodies to the ABA was shown. Total transcription in transcription system of cyanobacteria was activated by this ABA-binding proteins in vitro. This suggests, that some elements of the hormonal system of plants, which are associated with the ABA, was transfered from cyanobacterias to the eukaryotic cell.
Differential gene expression analysis (DESeq2) of Synechocystis culture treated with 20 mM of hex... more Differential gene expression analysis (DESeq2) of Synechocystis culture treated with 20 mM of hexan-1-ol for 30 min in 3 independent replications for each, "-" and "+" samples
Пленарные доклады 1Казанский институт биохимии и биофизики-обособленное структурное подразделение... more Пленарные доклады 1Казанский институт биохимии и биофизики-обособленное структурное подразделение ФИЦ КазНЦ РАН, Казань, Россия 2Казанский (Приволжский) федеральный университет, Казань, Россия, 3Федеральное государственное бюджетное учреждение науки Институт биохимии и физиологии растений и микроорганизмов РАН, Саратов, Россия, 4Институт органической и физической химии им. А.Е. Арбузова-обособленное структурное подразделение ФИЦ КазНЦ РАН, Казань, Россия
Клетки прокариот, в том числе и цианобактерий, отвечают на снижение температуры окружающей среды ... more Клетки прокариот, в том числе и цианобактерий, отвечают на снижение температуры окружающей среды активацией множества генов низкотемпературного ответа. Снижение температуры вызывает уменьшение текучести клеточных мембран, которая поддерживается на оптимальном уровне за счет активности десатураз жирных кислот (ЖК). Нами исследована температурозависимая экспрессия гена 3-десатуразы ЖК, desB, у цианобактерии Synechocystis, обладающей способностью синтезировать полиненасыщенные ЖК, а также у ее двойного мутанта, desA-/desD-, дефектного по генам 12- и -десатураз ЖК, для которого характерно наличие лишь мононенасыщенных ЖК. При снижении температуры в клетках дикого типа увеличивалось количество мРНК desB, достигая максимального значения при 24°С. У двойного мутанта desA-/desD- накопление транскрипта desB характеризовалось максимумом при 2830°С. Таким образом, на примере гена desB, кодирующего 3-десатуразу ЖК, показано, что температурозависимая экспрессия генов, отвечающих за поддержание оптимальной текучести клеточных мембран, определяется физическим состоянием этих мембран и регулируется по принципу обратной связи.
The main limiting factors for RNA-Seq analysis are quality and quantity of the isolated mRNA. In ... more The main limiting factors for RNA-Seq analysis are quality and quantity of the isolated mRNA. In prokaryotes, the proportion of messenger RNA to total RNA is rather low. Therefore, the main strategy of library preparation for sequencing is mRNA enrichment. Ribosomal and transfer RNAs, both monophosphorylated at the 5 0-ends, are the major fractions of total RNA, while the bulk of primary transcripts is triphosphorylated at the 5 0-teminus. Due to its low molecular weight, transfer RNA could be easily removed by a quick precipitation in LiCl solution. Ribosomal RNA may be degraded enzymatically by 5 0end terminal exonuclease XRN-1. These steps allow enriching samples in mRNA during the first stages of RNA-Seq library preparation. The desired level of fragmentation of enriched mRNA necessary for the 2 nd generation sequencing can be controlled by the duration of incubation at elevated temperatures in the presence of Mg 2þ-ions. Here, we describe a simple protocol for construction of the primary prokaryotic mRNA-saturated library without long depletion procedures.
Applied Biochemistry and Microbiology, Dec 1, 2017
⎯We isolated seven algologically and five bacteriologically pure cultures of toxin-producing cyan... more ⎯We isolated seven algologically and five bacteriologically pure cultures of toxin-producing cyanobacteria from Turgen gorge (Kazakhstan), Karlovy Vary (Czech Republic), and Shar-Nuur Lake, Bayan Ulgiiregion (Mongolia) springs. According to the Daphnia magna test, Desertifilum sp. and Nostoc sp. strains were the most toxic in the test of isolated strains (complete death of all test organisms was detected after 48 h). These strains possessed the highest inhibitory effect on proliferation of the HeLa cancer cell line. The Anabaena sp. 35 and Nostoc sp. 4 strains were also high toxic. Model strains Synechocystis PCC 6803 and Synechococcus elongates PCC 7942, as well as the strain isolated in the present work, Synechococcus sp. 55, were less toxic. Mass spectrometry made it possible to assign cyanobacterial toxins to cyclic depsipeptides. Two cyclic depsipeptides, micropeptin T and oscillapeptin, were detected in Desertifilum sp. extracts. Cryptophycin and small amounts of cyclic depsipeptide micropeptin SD were detected in Nostoc sp. extract.
Here, for the first time, we report the presence of highly active extracellular carbonic anhydras... more Here, for the first time, we report the presence of highly active extracellular carbonic anhydrase (CA) of α-class in cyanobacterial cells. The enzyme activity was confirmed both in vivo in intact cells and in vitro, using the recombinant protein. CA activity in intact cells of Cyanothece sp. ATCC 51142 reached ∼ 0.6 Wilbur-Anderson units (WAU) per 1 mg of total cell protein, and it was inhibited by a specific CAs inhibitor, ethoxyzolamide. The genes cce_4328 (ecaA) and cce_0871 (ecaB), encoding two potential extracellular CAs of Cyanothece have been cloned, and the corresponding proteins EcaA and EcaB, representing CAs of αand β-class, respectively, have been heterologously expressed in Escherichia coli. High specific activity (~1.1×10 4 WAU per 1 mg of target protein) was detected for the recombinant EcaA only. The presence of EcaA in the outer cellular layers of Cyanothece was confirmed by immunological analysis with antibodies raised against the recombinant protein. The absence of redox regulation of EcaA activity indicates that this protein does not possess a disulfide bond essential for some α-class CAs. The content and activity of EcaA in a fraction of periplasmic proteins was higher in Cyanothece cells grown at ambient concentration of CO 2 (0.04%) compared to those grown at an elevated CO 2 concentration (1.7%). At the same time, the level of ecaA gene mRNA varied insignificantly in response to changes in CO 2 supply. Our results indicate that EcaA is responsible for CA activity of intact Cyanothece cells and point to its possible physiological role of EcaA under low-CO 2 conditions. Highly active extracellular α α α α-class carbonic anhydrase of Cyanothece sp. ATCC 51142
Шар нуур көлінен токсин түзуші цианобактериялардың жаңа дақылдарын бөліп алу және идентификацияла... more Шар нуур көлінен токсин түзуші цианобактериялардың жаңа дақылдарын бөліп алу және идентификациялау Мақалада Моңғолияның Баян Өлгей аймағында орналасқан Шар Нуур (Сары көл) көлінің альгофлора құрамы зерттелді. Зерттелген су үлгілерінен Oscillatoriaceae және Nostocaceae тұқымдасының 3 бактериологиялық таза дақыл бөлініп алынды. Н.С. Строганов жүйесі бойынша бөлініп алынған дақылдардың токсинділігі анықталды. Тест-объект дафнияға қатынасы бойынша дақыл SP-O1 өте улы болды. Токсин түзуші дақылдың ісік жасуша HeLa тест-объектісіне қатынасын бағалау бойынша цитотоксинді белсенділік көрсетті. SP-O1 штамының экстрактісін зерттеу нәтижесі бойынша қауіпті токсиндер анықталған жоқ. Идентификацияланған токсиндер негізінен микроцистиндерге қарайды. Шар Нуур көлінен бөлініп алынған цианобактерия SP-O1 дақылы ботаникалық белгілері бойынша Oscillatoria туысына жатқызылғанмен, генетикалық сараптама бойынша олар Oscillatoriaceae тұқымдасының Desertifilum туысына жоғары гомологияны көрсетті. Осы мәліметтерге негізделе отырып SP-O1 дақылы идентификацияланып Desertifilum sp.1. деген атау берілді.
Пленарные доклады 1Казанский институт биохимии и биофизики-обособленное структурное подразделение... more Пленарные доклады 1Казанский институт биохимии и биофизики-обособленное структурное подразделение ФИЦ КазНЦ РАН, Казань, Россия 2Казанский (Приволжский) федеральный университет, Казань, Россия, 3Федеральное государственное бюджетное учреждение науки Институт биохимии и физиологии растений и микроорганизмов РАН, Саратов, Россия, 4Институт органической и физической химии им. А.Е. Арбузова-обособленное структурное подразделение ФИЦ КазНЦ РАН, Казань, Россия
Plants, algae, and photosynthetic bacteria experience frequent changes in environment. The abilit... more Plants, algae, and photosynthetic bacteria experience frequent changes in environment. The ability to survive depends on their capacity to acclimate to such changes. In particular, fluctuations in temperature affect the fluidity of cytoplasmic and thylakoid membranes. The molecular mechanisms responsible for the perception of changes in membrane fluidity have not been fully characterized. However, the understanding of the functions of the individual genes for fatty acid desaturases in cyanobacteria and plants led to the directed mutagenesis of such genes that altered the membrane fluidity of cytoplasmic and thylakoid membranes. Characterization of the photosynthetic properties of the transformed cyanobacteria and higher plants revealed that lipid unsaturation is essential for protection of the photosynthetic machinery against environmental stresses, such as strong light, salt stress, and high and low temperatures. The unsaturation of fatty acids enhances the repair of the damaged photosystem II complex under stress conditions. In this review, we summarize the knowledge on the mechanisms that regulate membrane fluidity, on putative sensors that perceive changes in membrane fluidity, on genes that are involved in acclimation to new sets of environmental conditions, and on the influence of membrane properties on photosynthetic functions.
... Transfer to Cyanobacteria in the Laboratory and in Nature 45 Enrique Flores, Alicia M ... 4 M... more ... Transfer to Cyanobacteria in the Laboratory and in Nature 45 Enrique Flores, Alicia M ... 4 Molecular Ecology and Environmental Genomics of Cyanobacteria 59 Ferran Garcia-Pichel 5 ... Symbioses: Signaling and Development 447 Birgitta Bergman, Liang Ran, and David G. Adams ...
Фенольные соединения относятся к веществам вторичного (специализированного) метаболизма, состав и... more Фенольные соединения относятся к веществам вторичного (специализированного) метаболизма, состав и содержание которых наиболее изучены в высших растениях, в отличие от прокариот. В настоящей работе представлены данные по изучению накопления этих метаболитов в процессе роста цианобактерий Synechocystis sp. (Synechocystis sp. PCC 6803 GT-L, штамм IPPAS В-1400) и Desertifilum tharense (штамм IPPAS B-1220) из коллекции микроводорослей и цианобактерий IPPAS ИФР РАН, а также исследованию их состава. Для этого были использованы методы спектрофотометрического анализа, тонкослойной хроматографии, УФ-спектрометрии. Установлено более высокое накопление фенольных соединений у Synechocystis sp., которое почти вдвое превышало таковое у D. tharense. В период линейной фазы роста (3 сутки) оно было выше такового в стационарную фазу роста (10 сутки) у обеих культур. Выявлены отличия в составе фенольных соединений, присутствующих в этанольных экстрактах Synechocystis sp. и D. tharense (4 и 7 веществ соответственно). Установлено наличие в них коньюгатов п-оксибензойной и п-кумаровой кислот − метаболитов начальных этапов биогенеза фенольных соединений.
Вестник Российского университета дружбы народов. Серия: Агрономия и животноводство, 2010
This article dedicates the study concerning evolution of the hormonal system of plants, in partic... more This article dedicates the study concerning evolution of the hormonal system of plants, in particular, detection of components of the regulatory system of abscisic acid (ABA) in cyanobacteria, which are considered as the precursors of the chloroplasts. ABA-binding proteins with molecular masses of 23, 50, 60 and 67 kDa were isolated using affinity chromatography on Sepharose conjugated with ABA from the lysate of cyanobacteria Synechocystis sp. PCC 6803. Interaction of isolated proteins with idiotypic antibodies to the ABA was shown. Total transcription in transcription system of cyanobacteria was activated by this ABA-binding proteins in vitro. This suggests, that some elements of the hormonal system of plants, which are associated with the ABA, was transfered from cyanobacterias to the eukaryotic cell.
Differential gene expression analysis (DESeq2) of Synechocystis culture treated with 20 mM of hex... more Differential gene expression analysis (DESeq2) of Synechocystis culture treated with 20 mM of hexan-1-ol for 30 min in 3 independent replications for each, "-" and "+" samples
Пленарные доклады 1Казанский институт биохимии и биофизики-обособленное структурное подразделение... more Пленарные доклады 1Казанский институт биохимии и биофизики-обособленное структурное подразделение ФИЦ КазНЦ РАН, Казань, Россия 2Казанский (Приволжский) федеральный университет, Казань, Россия, 3Федеральное государственное бюджетное учреждение науки Институт биохимии и физиологии растений и микроорганизмов РАН, Саратов, Россия, 4Институт органической и физической химии им. А.Е. Арбузова-обособленное структурное подразделение ФИЦ КазНЦ РАН, Казань, Россия
Клетки прокариот, в том числе и цианобактерий, отвечают на снижение температуры окружающей среды ... more Клетки прокариот, в том числе и цианобактерий, отвечают на снижение температуры окружающей среды активацией множества генов низкотемпературного ответа. Снижение температуры вызывает уменьшение текучести клеточных мембран, которая поддерживается на оптимальном уровне за счет активности десатураз жирных кислот (ЖК). Нами исследована температурозависимая экспрессия гена 3-десатуразы ЖК, desB, у цианобактерии Synechocystis, обладающей способностью синтезировать полиненасыщенные ЖК, а также у ее двойного мутанта, desA-/desD-, дефектного по генам 12- и -десатураз ЖК, для которого характерно наличие лишь мононенасыщенных ЖК. При снижении температуры в клетках дикого типа увеличивалось количество мРНК desB, достигая максимального значения при 24°С. У двойного мутанта desA-/desD- накопление транскрипта desB характеризовалось максимумом при 2830°С. Таким образом, на примере гена desB, кодирующего 3-десатуразу ЖК, показано, что температурозависимая экспрессия генов, отвечающих за поддержание оптимальной текучести клеточных мембран, определяется физическим состоянием этих мембран и регулируется по принципу обратной связи.
The main limiting factors for RNA-Seq analysis are quality and quantity of the isolated mRNA. In ... more The main limiting factors for RNA-Seq analysis are quality and quantity of the isolated mRNA. In prokaryotes, the proportion of messenger RNA to total RNA is rather low. Therefore, the main strategy of library preparation for sequencing is mRNA enrichment. Ribosomal and transfer RNAs, both monophosphorylated at the 5 0-ends, are the major fractions of total RNA, while the bulk of primary transcripts is triphosphorylated at the 5 0-teminus. Due to its low molecular weight, transfer RNA could be easily removed by a quick precipitation in LiCl solution. Ribosomal RNA may be degraded enzymatically by 5 0end terminal exonuclease XRN-1. These steps allow enriching samples in mRNA during the first stages of RNA-Seq library preparation. The desired level of fragmentation of enriched mRNA necessary for the 2 nd generation sequencing can be controlled by the duration of incubation at elevated temperatures in the presence of Mg 2þ-ions. Here, we describe a simple protocol for construction of the primary prokaryotic mRNA-saturated library without long depletion procedures.
Applied Biochemistry and Microbiology, Dec 1, 2017
⎯We isolated seven algologically and five bacteriologically pure cultures of toxin-producing cyan... more ⎯We isolated seven algologically and five bacteriologically pure cultures of toxin-producing cyanobacteria from Turgen gorge (Kazakhstan), Karlovy Vary (Czech Republic), and Shar-Nuur Lake, Bayan Ulgiiregion (Mongolia) springs. According to the Daphnia magna test, Desertifilum sp. and Nostoc sp. strains were the most toxic in the test of isolated strains (complete death of all test organisms was detected after 48 h). These strains possessed the highest inhibitory effect on proliferation of the HeLa cancer cell line. The Anabaena sp. 35 and Nostoc sp. 4 strains were also high toxic. Model strains Synechocystis PCC 6803 and Synechococcus elongates PCC 7942, as well as the strain isolated in the present work, Synechococcus sp. 55, were less toxic. Mass spectrometry made it possible to assign cyanobacterial toxins to cyclic depsipeptides. Two cyclic depsipeptides, micropeptin T and oscillapeptin, were detected in Desertifilum sp. extracts. Cryptophycin and small amounts of cyclic depsipeptide micropeptin SD were detected in Nostoc sp. extract.
Here, for the first time, we report the presence of highly active extracellular carbonic anhydras... more Here, for the first time, we report the presence of highly active extracellular carbonic anhydrase (CA) of α-class in cyanobacterial cells. The enzyme activity was confirmed both in vivo in intact cells and in vitro, using the recombinant protein. CA activity in intact cells of Cyanothece sp. ATCC 51142 reached ∼ 0.6 Wilbur-Anderson units (WAU) per 1 mg of total cell protein, and it was inhibited by a specific CAs inhibitor, ethoxyzolamide. The genes cce_4328 (ecaA) and cce_0871 (ecaB), encoding two potential extracellular CAs of Cyanothece have been cloned, and the corresponding proteins EcaA and EcaB, representing CAs of αand β-class, respectively, have been heterologously expressed in Escherichia coli. High specific activity (~1.1×10 4 WAU per 1 mg of target protein) was detected for the recombinant EcaA only. The presence of EcaA in the outer cellular layers of Cyanothece was confirmed by immunological analysis with antibodies raised against the recombinant protein. The absence of redox regulation of EcaA activity indicates that this protein does not possess a disulfide bond essential for some α-class CAs. The content and activity of EcaA in a fraction of periplasmic proteins was higher in Cyanothece cells grown at ambient concentration of CO 2 (0.04%) compared to those grown at an elevated CO 2 concentration (1.7%). At the same time, the level of ecaA gene mRNA varied insignificantly in response to changes in CO 2 supply. Our results indicate that EcaA is responsible for CA activity of intact Cyanothece cells and point to its possible physiological role of EcaA under low-CO 2 conditions. Highly active extracellular α α α α-class carbonic anhydrase of Cyanothece sp. ATCC 51142
Шар нуур көлінен токсин түзуші цианобактериялардың жаңа дақылдарын бөліп алу және идентификацияла... more Шар нуур көлінен токсин түзуші цианобактериялардың жаңа дақылдарын бөліп алу және идентификациялау Мақалада Моңғолияның Баян Өлгей аймағында орналасқан Шар Нуур (Сары көл) көлінің альгофлора құрамы зерттелді. Зерттелген су үлгілерінен Oscillatoriaceae және Nostocaceae тұқымдасының 3 бактериологиялық таза дақыл бөлініп алынды. Н.С. Строганов жүйесі бойынша бөлініп алынған дақылдардың токсинділігі анықталды. Тест-объект дафнияға қатынасы бойынша дақыл SP-O1 өте улы болды. Токсин түзуші дақылдың ісік жасуша HeLa тест-объектісіне қатынасын бағалау бойынша цитотоксинді белсенділік көрсетті. SP-O1 штамының экстрактісін зерттеу нәтижесі бойынша қауіпті токсиндер анықталған жоқ. Идентификацияланған токсиндер негізінен микроцистиндерге қарайды. Шар Нуур көлінен бөлініп алынған цианобактерия SP-O1 дақылы ботаникалық белгілері бойынша Oscillatoria туысына жатқызылғанмен, генетикалық сараптама бойынша олар Oscillatoriaceae тұқымдасының Desertifilum туысына жоғары гомологияны көрсетті. Осы мәліметтерге негізделе отырып SP-O1 дақылы идентификацияланып Desertifilum sp.1. деген атау берілді.
Пленарные доклады 1Казанский институт биохимии и биофизики-обособленное структурное подразделение... more Пленарные доклады 1Казанский институт биохимии и биофизики-обособленное структурное подразделение ФИЦ КазНЦ РАН, Казань, Россия 2Казанский (Приволжский) федеральный университет, Казань, Россия, 3Федеральное государственное бюджетное учреждение науки Институт биохимии и физиологии растений и микроорганизмов РАН, Саратов, Россия, 4Институт органической и физической химии им. А.Е. Арбузова-обособленное структурное подразделение ФИЦ КазНЦ РАН, Казань, Россия
Plants, algae, and photosynthetic bacteria experience frequent changes in environment. The abilit... more Plants, algae, and photosynthetic bacteria experience frequent changes in environment. The ability to survive depends on their capacity to acclimate to such changes. In particular, fluctuations in temperature affect the fluidity of cytoplasmic and thylakoid membranes. The molecular mechanisms responsible for the perception of changes in membrane fluidity have not been fully characterized. However, the understanding of the functions of the individual genes for fatty acid desaturases in cyanobacteria and plants led to the directed mutagenesis of such genes that altered the membrane fluidity of cytoplasmic and thylakoid membranes. Characterization of the photosynthetic properties of the transformed cyanobacteria and higher plants revealed that lipid unsaturation is essential for protection of the photosynthetic machinery against environmental stresses, such as strong light, salt stress, and high and low temperatures. The unsaturation of fatty acids enhances the repair of the damaged photosystem II complex under stress conditions. In this review, we summarize the knowledge on the mechanisms that regulate membrane fluidity, on putative sensors that perceive changes in membrane fluidity, on genes that are involved in acclimation to new sets of environmental conditions, and on the influence of membrane properties on photosynthetic functions.
... Transfer to Cyanobacteria in the Laboratory and in Nature 45 Enrique Flores, Alicia M ... 4 M... more ... Transfer to Cyanobacteria in the Laboratory and in Nature 45 Enrique Flores, Alicia M ... 4 Molecular Ecology and Environmental Genomics of Cyanobacteria 59 Ferran Garcia-Pichel 5 ... Symbioses: Signaling and Development 447 Birgitta Bergman, Liang Ran, and David G. Adams ...
Uploads
Papers by Dmitry Los