scholarly journals pH-dependent regulation of the multi-subunit cation/proton antiporter Pha1 system from Sinorhizobium meliloti

Microbiology ◽  
2009 ◽  
Vol 155 (8) ◽  
pp. 2750-2756 ◽  
Author(s):  
Toshio Yamaguchi ◽  
Fuminori Tsutsumi ◽  
Péter Putnoky ◽  
Masahiro Fukuhara ◽  
Tatsunosuke Nakamura
Keyword(s):  
Sinorhizobium Meliloti ◽  
Sequence Similarity ◽  
Plant Root ◽  
Functional Expression ◽  
Amino Acid Sequence Similarity ◽  
Ph Optimum ◽  
Cation Selectivity ◽  
High Amino Acid ◽  
Alkaline Conditions ◽  
Ph Dependent

The pha1 gene cluster (pha1A′-G) of Sinorhizobium meliloti has previously been characterized as a necessary component for proper invasion into plant root tissue. It has been suggested to encode a multi-subunit K+/H+ antiporter, since mutations in the pha1 region rendered S. meliloti cells sensitive to K+ and alkali, and because there is high amino acid sequence similarity to previously characterized multi-subunit cation/H+ antiporters (Mrp antiporters). However, the detailed transport properties of the Pha1 system are yet to be determined. Interestingly, most of the Mrp antiporters are highly selective for Na+, unlike the Pha1 system. Here, we report the functional expression of the Pha1 system in Escherichia coli and the measurement of cation/H+ antiport activity. We showed that the Pha1 system is indeed a K+/H+ antiporter with a pH optimum under mildly alkaline conditions. Moreover, we found that the Pha1 system can transport Na+; this was unexpected based on previous phenotypic analyses of pha1 mutants. Furthermore, we demonstrated that the cation selectivity of the Pha1 system was altered when the pH was lowered from the optimum. The downregulation of Na+/H+ and K+/H+ antiport activities upon acidic shift appeared to occur via different processes, which might indicate the presence of distinct mechanisms for the regulation of the K+/H+ and Na+/H+ antiport activities of the Pha1 system.

scholarly journals Identification and Characterization of a NaCl-Responsive Genetic Locus Involved in Survival during Desiccation in Sinorhizobium meliloti

2013 ◽  
Vol 79 (18) ◽  
pp. 5693-5700 ◽  
Author(s):  
Jan A. C. Vriezen ◽  
Frans J. de Bruijn ◽  
Klaus Nüsslein
Keyword(s):  
Sinorhizobium Meliloti ◽  
Agricultural Soils ◽  
Sequence Similarity ◽  
Genetic Locus ◽  
Model Organism ◽  
Normal Growth ◽  
Growth Conditions ◽  
Amino Acid Sequence Similarity ◽  
Content Type ◽  
Genetic Mechanisms

ABSTRACTTheRhizobiaceaeare a bacterial family of enormous agricultural importance due to the ability of its members to fix atmospheric nitrogen in an intimate relationship with plants. Their survival as naturally occurring soil bacteria in agricultural soils as well as popular seed inocula is affected directly by drought and salinity. Survival after desiccation in the presence of NaCl is enabled by underlying genetic mechanisms in the model organismSinorhizobium meliloti1021. Since salt stress parallels a loss in water activity, the identification of NaCl-responsive loci may identify loci involved in survival during desiccation. This approach enabled identification of the lociasnOandnggby their reduced ability to grow on increased NaCl concentrations, likely due to their inability to produce the osmoprotectant N-acetylglutaminylglutamine (NAGGN). In addition, the mutant harboringngg::Tn5luxABwas affected in its ability to survive desiccation and responded to osmotic stress. The desiccation sensitivity may have been due to secondary functions of Ngg (N-acetylglutaminylglutamine synthetase)-like cell wall metabolism as suggested by the presence of ad-alanine-d-alanine ligase (dAla-dAla) domain and by sensitivity of the mutant to β-lactam antibiotics.asnO::Tn5luxABis expressed during the stationary phase under normal growth conditions. Amino acid sequence similarity to enzymes producing β-lactam inhibitors and increased resistance to β-lactam antibiotics may indicate thatasnOis involved in the production of a β-lactam inhibitor.

scholarly journals Inhibition of mitogen-activated protein kinase by a Drosophila dual-specific phosphatase

2000 ◽  
Vol 349 (3) ◽  
pp. 821-828 ◽  
Author(s):  
Won-Jae LEE ◽  
Sun-Hong KIM ◽  
Yong-Sik KIM ◽  
Sung-Jun HAN ◽  
Ki-Sook PARK ◽  
...  
Keyword(s):  
Amino Acid ◽  
Protein Kinase ◽  
Expressed Sequence Tag ◽  
Mapk Pathway ◽  
Sequence Similarity ◽  
Mitogen Activated Protein Kinase ◽  
Amino Acid Sequence Similarity ◽  
Calculated Molecular Mass ◽  
High Amino Acid ◽  
Mitogen Activated Protein

The Drosophila extracellular signal-regulated kinase (DERK) mitogen-activated protein kinase (MAPK) is involved in the regulation of multiple differentiation and developmental processes. Tight control of MAPK activity is critical for normal cell behaviour. We identified a novel Drosophila MAPK phosphatase (DMKP) cDNA from the expressed-sequence-tag database and characterized it. Analysis of the nucleotide sequence revealed an open reading frame encoding the 203-amino acid protein, with a calculated molecular mass of 23kDa, which has a high amino acid sequence similarity with ‘VH1-like’dual-specific phosphatases at the broad region near the catalytic sites. The expression of DMKP mRNA occurs from the late larval stages to adulthood in Drosophila development. The recombinant DMKP protein produced in yeast retained its phosphatase activity. When expressed in Schneider cells, DMKP dose-dependently inhibited DERK and Drosophila c-Jun N-terminal kinase activities with high selectivity towards DERK. However, DMKP did not have any affect on Drosophila p38 activity. When DMKP was expressed in yeast, it down-regulated the fus1-lacZ trans-reporter gene of the pheromone MAPK pathway without any significant effect on the high-osmolarity-glycerol-response pathway.

scholarly journals Cloning of the Gene Encoding a Novel Thermostable α-Galactosidase from Thermus brockianus ITI360

1999 ◽  
Vol 65 (9) ◽  
pp. 3955-3963 ◽  
Author(s):  
Olafur Fridjonsson ◽  
Hildegard Watzlawick ◽  
Axel Gehweiler ◽  
Thilo Rohrhirsch ◽  
Ralf Mattes
Keyword(s):  
Thermotoga Maritima ◽  
Sequence Similarity ◽  
Amino Acid Sequence Similarity ◽  
Thermotoga Neapolitana ◽  
Glycosyl Hydrolases ◽  
Gene Encoding ◽  
Ph Optimum ◽  
Carbohydrate Source ◽  
Calculated Molecular Mass

ABSTRACT An α-galactosidase gene from Thermus brockianusITI360 was cloned, sequenced, and expressed in Escherichia coli, and the recombinant protein was purified. The gene, designated agaT, codes for a 476-residue polypeptide with a calculated molecular mass of 53,810 Da. The native structure of the recombinant enzyme (AgaT) was estimated to be a tetramer. AgaT displays amino acid sequence similarity to the α-galactosidases ofThermotoga neapolitana and Thermotoga maritimaand a low-level sequence similarity to α-galactosidases of family 36 in the classification of glycosyl hydrolases. The enzyme is thermostable, with a temperature optimum of activity at 93°C withpara-nitrophenyl-α-galactopyranoside as a substrate. Half-lives of inactivation at 92 and 80°C are 100 min and 17 h, respectively. The pH optimum is between 5.5 and 6.5. The enzyme displayed high affinity for oligomeric substrates. TheKm s for melibiose and raffinose at 80°C were determined as 4.1 and 11.0 mM, respectively. The α-galactosidase gene in T. brockianus ITI360 was inactivated by integrational mutagenesis. Consequently, no α-galactosidase activity was detectable in crude extracts of the mutant strain, and it was unable to use melibiose or raffinose as a single carbohydrate source.

scholarly journals The Trichohyalin-Like Protein Scaffoldin Is Expressed in the Multilayered Periderm during Development of Avian Beak and Egg Tooth

Genes ◽  
2021 ◽  
Vol 12 (2) ◽  
pp. 248
Author(s):  
Veronika Mlitz ◽  
Marcela Hermann ◽  
Maria Buchberger ◽  
Erwin Tschachler ◽  
Leopold Eckhart
Keyword(s):  
Sequence Similarity ◽  
Coturnix Japonica ◽  
Amino Acid Sequence Similarity ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Embryonic Tissues ◽  
Lower Beak ◽  
High Amino Acid ◽  
Evolutionarily Conserved ◽  
Polymerase Chain

Scaffoldin, an S100 fused-type protein (SFTP) with high amino acid sequence similarity to the mammalian hair follicle protein trichohyalin, has been identified in reptiles and birds, but its functions are not yet fully understood. Here, we investigated the expression pattern of scaffoldin and cornulin, a related SFTP, in the developing beaks of birds. We determined the mRNA levels of both SFTPs by reverse transcription polymerase chain reaction (RT-PCR) in the beak and other ectodermal tissues of chicken (Gallus gallus) and quail (Coturnix japonica) embryos. Immunohistochemical staining was performed to localize scaffoldin in tissues. Scaffoldin and cornulin were expressed in the beak and, at lower levels, in other embryonic tissues of both chickens and quails. Immunohistochemistry revealed scaffoldin in the peridermal compartment of the egg tooth, a transitory cornified protuberance (caruncle) on the upper beak which breaks the eggshell during hatching. Furthermore, scaffoldin marked a multilayered peridermal structure on the lower beak. The results of this study suggest that scaffoldin plays an evolutionarily conserved role in the development of the avian beak with a particular function in the morphogenesis of the egg tooth.

425. Expression of mammalian cysteine-rich secretory proteins in the mouse model

2008 ◽  
Vol 20 (9) ◽  
pp. 105
Author(s):  
T. Reddy ◽  
G. M. Gibbs ◽  
D. J. Merriner ◽  
J. B. Kerr ◽  
M. K. O.'Bryan
Keyword(s):  
Expression Profile ◽  
Reproductive Tract ◽  
Sequence Similarity ◽  
Immunohistochemical Analysis ◽  
The Body ◽  
Mouse Tissue ◽  
Amino Acid Sequence Similarity ◽  
Secretory Proteins ◽  
Male Reproductive Tract ◽  
High Amino Acid

Mammalian cysteine rich secretory proteins are a family of four proteins exhibiting a high amino acid sequence similarity and belonging to the CAP (Cysteine rich secretory proteins, Antigen-5 proteins and the plant Pathogenesis related-1 proteins) superfamily of proteins. They are designated CRISP 1, 2, 3 and 4. Structurally, mammalian CRISP’s are characterised by 16 cysteine residues involved in intra-molecular di-sulfide bonds and the formation of 2 domains, ie., the CRISP domain (CD) and CAP domain. Whilst studies on mouse CRISP2 suggest that the CD is involved in ion channel regulation, studies on non-mammalian CAP superfamily members suggest that the CAP domain is involved in proteolytic activity.They are predominantly expressed and localised in the male reproductive tract, however, the EST expression databases suggest that mammalian CRISPs are expressed more widely than in the male reproductive tract. The objective of this study was therefore to conclusively define the expression and localisation of each CRISP protein in a mammalian system.A reverse transcription PCR expression profile and immunohistochemical analysis of 16 mouse tissue was conducted to establish the expression and localisation of each of the four CRISPs. These data showed that although the CRISPs have a strong expression and localisation bias to the male reproductive tract, they are widely distributed throughout the body in mice, including the ovary, uterus, and mammary gland. Whilst each CRISP has a clear expression profile, there was a striking localisation of androgen regulated CRISPs (1, 3, 4) in immune tissue including the spleen and thymus. Such a localisation raises the spectre of a role for CRISPs in the normal physiology and disease of several organs.

scholarly journals NrdH Redoxin Enhances Resistance to Multiple Oxidative Stresses by Acting as a Peroxidase Cofactor in Corynebacterium glutamicum

2013 ◽  
Vol 80 (5) ◽  
pp. 1750-1762 ◽  
Author(s):  
Mei-Ru Si ◽  
Lei Zhang ◽  
Zhi-Fang Yang ◽  
Yi-Xiang Xu ◽  
Ying-Bao Liu ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Corynebacterium Glutamicum ◽  
Sequence Similarity ◽  
Thioredoxin Reductase ◽  
Amino Acid Sequence Similarity ◽  
Content Type ◽  
Reduction Activity ◽  
Oxidative Stresses ◽  
High Amino Acid ◽  
Thiol Peroxidase

ABSTRACTNrdH redoxins are small protein disulfide oxidoreductases behaving like thioredoxins but sharing a high amino acid sequence similarity to glutaredoxins. Although NrdH redoxins are supposed to be another candidate in the antioxidant system, their physiological roles in oxidative stress remain unclear. In this study, we confirmed that theCorynebacterium glutamicumNrdH redoxin catalytically reduces the disulfides in the class Ib ribonucleotide reductases (RNR), insulin and 5,5′-dithiobis-(2-nitrobenzoic acid) (DTNB), by exclusively receiving electrons from thioredoxin reductase. Overexpression of NrdH increased the resistance ofC. glutamicumto multiple oxidative stresses by reducing ROS accumulation. Accordingly, elevated expression of thenrdHgene was observed when theC. glutamicumwild-type strain was exposed to oxidative stress conditions. It was discovered that the NrdH-mediated resistance to oxidative stresses was largely dependent on the presence of the thiol peroxidase Prx, as the increased resistance to oxidative stresses mediated by overexpression of NrdH was largely abrogated in theprxmutant. Furthermore, we showed that NrdH facilitated the hydroperoxide reduction activity of Prx by directly targeting and serving as its electron donor. Thus, we present evidence that the NrdH redoxin can protect against the damaging effects of reactive oxygen species (ROS) induced by various exogenous oxidative stresses by acting as a peroxidase cofactor.

scholarly journals A New d,l-Endopeptidase Gene Product, YojL (Renamed CwlS), Plays a Role in Cell Separation with LytE and LytF in Bacillus subtilis

2006 ◽  
Vol 188 (15) ◽  
pp. 5541-5550 ◽  
Author(s):  
Tatsuya Fukushima ◽  
Anahita Afkham ◽  
Shin-ichirou Kurosawa ◽  
Taichi Tanabe ◽  
Hiroki Yamamoto ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bacillus Subtilis ◽  
Cell Separation ◽  
Sequence Similarity ◽  
Terminal Region ◽  
Diaminopimelic Acid ◽  
Amino Acid Sequence Similarity ◽  
Northern Hybridization ◽  
Triple Mutant ◽  
High Amino Acid

ABSTRACT A new peptidoglycan hydrolase, Bacillus subtilis YojL (cell wall-lytic enzyme associated with cell separation, renamed CwlS), exhibits high amino acid sequence similarity to LytE (CwlF) and LytF (CwlE), which are associated with cell separation. The N-terminal region of CwlS has four tandem repeat regions (LysM repeats) predicted to be a peptidoglycan-binding module. The C-terminal region exhibits high similarity to the cell wall hydrolase domains of LytE and LytF at their C-terminal ends. The C-terminal region of CwlS produced in Escherichia coli could hydrolyze the linkage of d-γ-glutamyl-meso-diaminopimelic acid in the cell wall of B. subtilis, suggesting that CwlS is a d,l-endopeptidase. β-Galactosidase fusion experiments and Northern hybridization analysis suggested that the cwlS gene is transcribed during the late vegetative and early stationary phases. A cwlS mutant exhibited a cell shape similar to that of the wild type; however, a lytE lytF cwlS triple mutant exhibited aggregated microfiber formation. Moreover, immunofluorescence microscopy showed that FLAG-tagged CwlS was localized at cell separation sites and cell poles during the late vegetative phase. The localization sites are similar to those of LytF and LytE, indicating that CwlS is involved in cell separation with LytF and LytE. These specific localizations may be dependent on the LysM repeats in their N-terminal domains. The roles of CwlS, LytF, and LytE in cell separation are discussed.

scholarly journals Identification and Characterization of a Novel Intracellular Alkaline α-Amylase from the Hyperthermophilic Bacterium Thermotoga maritima MSB8

2006 ◽  
Vol 72 (3) ◽  
pp. 2206-2211 ◽  
Author(s):  
Meike Ballschmiter ◽  
Ole Fütterer ◽  
Wolfgang Liebl
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Thermotoga Maritima ◽  
Sequence Similarity ◽  
Hydrolytic Activity ◽  
Amino Acid Sequence Similarity ◽  
Glycoside Hydrolase Family ◽  
Ph Optimum ◽  
Intracellular Enzyme ◽  
Hyperthermophilic Bacterium

ABSTRACT The gene for a novel α-amylase, designated AmyC, from the hyperthermophilic bacterium Thermotoga maritima was cloned and heterologously overexpressed in Escherichia coli. The putative intracellular enzyme had no amino acid sequence similarity to glycoside hydrolase family (GHF) 13 α-amylases, yet the range of substrate hydrolysis and the product profile clearly define the protein as an α-amylase. Based on sequence similarity AmyC belongs to a subgroup within GHF 57. On the basis of amino acid sequence similarity, Glu185 and Asp349 could be identified as the catalytic residues of AmyC. Using a 60-min assay, the maximum hydrolytic activity of the purified enzyme, which was dithiothreitol dependent, was found to be at 90°C. AmyC displayed a remarkably high pH optimum of pH 8.5 and an unusual sensitivity towards both ATP and EDTA.

scholarly journals A tuber mustard AP2/ERF transcription factor gene, BjABR1, functioning in abscisic acid and abiotic stress responses, and evolutionary trajectory of the ABR1 homologous genes in Brassica species

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e6071 ◽  
Author(s):  
Liuxin Xiang ◽  
Chao Liu ◽  
Jingzhi Luo ◽  
Lin He ◽  
Yushan Deng ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Abscisic Acid ◽  
Abiotic Stress ◽  
Stress Responses ◽  
Sequence Similarity ◽  
Amino Acid Sequence Similarity ◽  
Genome Database ◽  
Homologous Genes ◽  
High Amino Acid ◽  
Tuber Mustard

The AP2/ERF superfamily of transcription factors is one of the largest transcription factor families in plants and plays an important role in plant development processes and stress responses. In this study, BjABR1, an AP2/ERF superfamily gene, from tuber mustard (Brassica juncea var. tumida Tsen et Lee), sharing high amino acid sequence similarity with the AtABR1 (Arabidopsis thaliana AP2-like abscisic acid repressor 1) gene, were performed functional research, and the ABR1 homologous genes in Brassica species were identified and performed phylogenetic analysis. The promoter sequence of BjABR1 contained many phytohormone- and stress-related cis-elements; ABA (abscisic acid) and abiotic stresses can induce BjABR1 expression in tuber mustard; overexpression of BjABR1 in Arabidopsis can alleviate plant sensitivity to ABA and salt and osmotic stresses, and the alleviation may be due to changes in stress/ABA-induced gene expression. These results indicated that BjABR1 functions in ABA and abiotic stress responses. By BLAST searches against the genome database of five Brassica species (three diploids, B. rapa, B. nigra, and B. oleracea, and two allotetraploid, B. juncea and B. napus) using the protein sequence of AtABR1, 3, 3, 3, 6, and 5 ABR1 homologous genes in B. nigra, B. rapa, B. oleracea, B. juncea, and B. napus were identified, respectively, and they shared high sequence similarity. By sequence analysis, annotation mistakes of the protein-coding regions of two ABR1 homologous genes, GSBRNA2T00134741001 and BjuB007684, were found and corrected. Then, the evolution analysis of these ABR1 homologous genes showed that the ancestor of the three diploid species had three ABR1 homologous genes and each diploid inherited all the three genes from their ancestor; then, allotetraploid B. juncea inherited all the six genes from B. rapa and B. nigra with no gene lost, while allotetraploid B. napus inherited all the three genes from B. oleracea and two genes from B. rapa with one gene lost, indicating that ABR1 homologous genes possessed greater hereditary conservation in Brassica species. The ABR1 homologous genes between B. rapa and B. oleracea shared much higher sequence similarity compared to that of B. nigra in diploid species, indicating that ABR1 homologous genes in B. nigra had experienced more rapid evolution, and B. rapa and B. oleracea may share closer relationship compared to B. nigra. Moreover, the spatial and temporal expression analysis of six ABR1 homologous genes of tuber mustard showed that they possessed different expression models. These results imply that ABR1 homologous genes are important to Brassica plants, and they may possess similar function in ABA and abiotic stress responses but play a role in different tissues and growing stages of plant. This study will provide the foundation to the functional research of ABR1 homologous genes in the Brassica species and help to reveal and understand the evolution mechanisms of Brassica species.

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