scholarly journals Analysis of a Symbiosis-Specific Cytochrome P450 Homolog in Rhizobium sp. BR816

2001 ◽  
Vol 14 (7) ◽  
pp. 918-924 ◽  
Author(s):  
Ellen Luyten ◽  
Elfriede Swinnen ◽  
Katrien Vlassak ◽  
Christel Verreth ◽  
Bruno Dombrecht ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Amino Acid ◽  
Transcriptional Analysis ◽  
Free Living ◽  
Amino Acid Similarity ◽  
Reading Frame ◽  
P450 Gene ◽  
Symbiotic Plasmid ◽  
Rhizobium Sp ◽  
Free Living Conditions

Sequence analysis of the DNA region upstream of nodO in Rhizobium sp. BR816 revealed an open reading frame in which the deduced amino acid sequence shows homology with cytochrome P450. Because the BR816 P450 homolog shows 73% amino acid similarity with CYP127A1(Y4vG), which is identified on the symbiotic plasmid of Rhizobium sp. NGR234, it is named CYP127A2. Transcriptional analysis of CYP127A2 revealed high expression in bacteroids, whereas no or hardly any expression was observed under free-living conditions. Low-level, free-living expression, however was noticed when cells were grown microoxically at acid pH levels. A number of possible substrates that may induce P450 gene expression were analyzed, but only the addition of short-chain alcohols to cultures slightly increased CYP127A2 expression. High levels of CYP127A2 expression observed in bacteroids of a nifH mutant strain, which formed non-fixing nodules on bean, indicated that the genuine substrate for CYP127A2 is not a metabolite resulting from N2-fixation. Nevertheless, expression analysis pointed to a NifA- and σ54-dependent transcription.

Plasmid replicons of Rhizobium

2005 ◽  
Vol 33 (1) ◽  
pp. 157-158 ◽  
Author(s):  
L.C. Crossman
Keyword(s):  
Short Review ◽  
Nitrogen Fixing ◽  
Rhizobium Etli ◽  
Free Living ◽  
Leguminous Plants ◽  
Symbiotic Plasmid ◽  
Rhizobium Spp ◽  
Plasmid Replicons ◽  
Rhizobium Sp

Rhizobium spp. are found in soil. They are both free-living and found symbiotically associated with the nodules of leguminous plants. Traditionally, studies have focused on the association of these organisms with plants in nitrogen-fixing nodules, since this is regarded as the most important role of these bacteria in the environment. Rhizobium sp. are known to possess several replicons. Some, like the Rhizobium etli symbiotic plasmid p42d and the plasmid pNGR234b of Rhizobium NGR234, have been sequenced and characterized. The plasmids from these organisms are the focus of this short review.

scholarly journals Cloning of a Wound Inducible Lycopersicon esculentum Cytochrome P450 Gene and Lack of Regeneration of Transgenic Plants with Sense or Antisense Constructs

2002 ◽  
Vol 127 (4) ◽  
pp. 535-539 ◽  
Author(s):  
Grzegorz Bartoszewski ◽  
Cesar V. Mujer ◽  
Katarzyna Niemirowicz-Szczytt ◽  
Ann C. Smigocki
Keyword(s):  
Cytochrome P450 ◽  
Transgenic Plants ◽  
Lycopersicon Esculentum ◽  
Nicotiana Plumbaginifolia ◽  
Mechanical Wounding ◽  
Cytochrome P450 Gene ◽  
Transcript Levels ◽  
Reading Frame ◽  
P450 Gene ◽  
Young Leaves

A Lycopersicon esculentum Mill. (tomato) cDNA clone with high similarity to a Nicotiana plumbaginifolia Viv. (tobacco) cytochrome P450 gene was isolated using 5' and 3' rapid amplification of cDNA ends (RACE). The isolated cDNA (GenBank Accession No. AF249329) has an open reading frame of 1494 base pairs (bp) and encodes a protein of 498 amino acids with 75% identity to the N. plumbaginifolia cytochrome P450 (CYP72A2) and 45% to a Catharanthus roseus G. Don (Madagaskar periwinkle) CYP72A1 protein sequence. By Southern-blot analysis, one or two highly homologous genes were detected in the L. esculentum genome. Expression of the cloned P450 gene was regulated by circadian rhythm and enhanced by wounding. Leaf transcripts were detected in the light but not dark. Highest transcript levels were observed 3 hours after mechanical wounding. No increase in expression was seen in response to applications of zeatin as with the N. plumbaginifolia gene. Of the tissues analyzed, shoot tips and young leaves and fruit had the highest detectable transcript levels. Attempts to transform more than 1400 cotyledon explants of L. esculentum with sense or antisense CYP72A2 gene constructs produced no transgenic plants.

scholarly journals nifDK Clusters Located on the Chromosome and Megaplasmid of Bradyrhizobium sp. Strain DOA9 Contribute Differently to Nitrogenase Activity During Symbiosis and Free-Living Growth

2016 ◽  
Vol 29 (10) ◽  
pp. 767-773 ◽  
Author(s):  
Jenjira Wongdee ◽  
Pongpan Songwattana ◽  
Nico Nouwen ◽  
Rujirek Noisangiam ◽  
Joel Fardoux ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Nitrogen Fixation ◽  
Nitrogenase Activity ◽  
Transcriptional Analysis ◽  
Main Role ◽  
Free Living ◽  
Living State ◽  
Bradyrhizobium Sp ◽  
Reporter Strains ◽  
Free Living Conditions

Bradyrhizobium sp. strain DOA9 contains two copies of the nifDK genes, nifDKc, located on the chromosome, and nifDKp, located on a symbiotic megaplasmid. Unlike most rhizobia, this bacterium displays nitrogenase activity under both free-living and symbiotic conditions. Transcriptional analysis using gusA reporter strains showed that both nifDK operons were highly expressed under symbiosis, whereas nifDKc was the most abundantly expressed under free-living conditions. During free-living growth, the nifDKp mutation did not affect nitrogenase activity, whereas nitrogenase activity was drastically reduced with the nifDKc mutant. This led us to suppose that nifDKc is the main contributor of nitrogenase activity in the free-living state. In contrast, during symbiosis, no effect of the nifDKc mutation was observed and the nitrogen-fixation efficiency of plants inoculated with the nifDKp mutant was reduced. This suggests that nifDKp plays the main role in nitrogenase enzyme activity during symbiosis. Together, these data suggest that Bradyrhizobium sp. strain DOA9 contains two functional copies of nifDK genes that are regulated differently and that, depending on their lifestyle, contribute differently to nitrogenase activity.

scholarly journals Identification, Characterization, and Azole-Binding Properties of Mycobacterium smegmatis CYP164A2, a Homolog of ML2088, the Sole Cytochrome P450 Gene of Mycobacterium leprae

2008 ◽  
Vol 53 (3) ◽  
pp. 1157-1164 ◽  
Author(s):  
Andrew G. S. Warrilow ◽  
Colin J. Jackson ◽  
Josie E. Parker ◽  
Timothy H. Marczylo ◽  
Diane E. Kelly ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Spin State ◽  
Mycobacterium Smegmatis ◽  
Spectral Characteristics ◽  
Mycobacterium Leprae ◽  
Cytochrome P450 Monooxygenase ◽  
P450 Monooxygenase ◽  
Reading Frame ◽  
Binding Characteristics ◽  
P450 Gene

ABSTRACT The genome sequence of Mycobacterium leprae revealed a single open reading frame, ML2088 (CYP164A1), encoding a putative full-length cytochrome P450 monooxygenase and 12 pseudogenes. We have identified a homolog of ML2088 in Mycobacterium smegmatis and report here the cloning, expression, purification, and azole-binding characteristics of this cytochrome P450 (CYP164A2). CYP164A2 is 1,245 bp long and encodes a protein of 414 amino acids and molecular mass of 45 kDa. CYP164A2 has 60% identity with Mycobacterium leprae CYP161A1 and 66 to 69% identity with eight other mycobacterial CYP164A1 homologs, with three identified highly conserved motifs. Recombinant CYP164A2 has the typical spectral characteristics of a cytochrome P450 monooxygenase, predominantly in the ferric low-spin state. Unusually, the spin state was readily modulated by increasing ionic strength at pH 7.5, with 50% high-spin occupancy achieved with 0.14 M NaCl. CYP164A2 bound clotrimazole, econazole, and miconazole strongly (Kd , 1.2 to 2.5 μM); however, strong binding with itraconazole, ketoconazole, and voriconazole was only observed in the presence of 0.5 M NaCl. Fluconazole did not bind to CYP164A2 at pH 7.5 and no discernible type II binding spectrum was observed.

scholarly journals High-resolution transcriptional analysis of the symbiotic plasmid of Rhizobium sp. NGR234

1999 ◽  
Vol 32 (2) ◽  
pp. 415-425 ◽  
Author(s):  
Xavier Perret ◽  
Christoph Freiberg ◽  
Andre Rosenthal ◽  
William J. Broughton ◽  
Remy Fellay
Keyword(s):  
High Resolution ◽  
Transcriptional Analysis ◽  
Symbiotic Plasmid ◽  
Rhizobium Sp

Gene Cloning, Transcriptional Analysis, Purification, and Characterization of Phenolic Acid Decarboxylase from Bacillus subtilis

1998 ◽  
Vol 64 (4) ◽  
pp. 1466-1471 ◽  
Author(s):  
Jean-François Cavin ◽  
Véronique Dartois ◽  
Charles Diviès
Keyword(s):  
Bacillus Subtilis ◽  
Amino Acid ◽  
Substrate Specificity ◽  
Phenolic Acid ◽  
Genomic Library ◽  
Transcriptional Analysis ◽  
Acid Decarboxylase ◽  
Reading Frame ◽  
Appl Environ ◽  
Phenolic Acid Decarboxylase

ABSTRACT Bacillus subtilis displays a substrate-inducible decarboxylating activity with the following three phenolic acids: ferulic, p-coumaric, and caffeic acids. Based on DNA sequence homologies between the Bacillus pumilus ferulate decarboxylase gene (fdc) (A. Zago, G. Degrassi, and C. V. Bruschi, Appl. Environ. Microbiol. 61:4484–4486, 1995) and theLactobacillus plantarum p-coumarate decarboxylase gene (pdc) (J.-F. Cavin, L. Barthelmebs, and C. Diviès, Appl. Environ. Microbiol. 63:1939–1944, 1997), a DNA probe of about 300 nucleotides for the L. plantarum pdcgene was used to screen a B. subtilis genomic library in order to clone the corresponding gene in this bacterium. One clone was detected with this heterologous probe, and this clone exhibited phenolic acid decarboxylase (PAD) activity. The corresponding 5-kb insertion was partially sequenced and was found to contain a 528-bp open reading frame coding for a 161-amino-acid protein exhibiting 71 and 84% identity with the pdc- and fdc-encoded enzymes, respectively. The PAD gene (pad) is transcriptionally regulated by p-coumaric, ferulic, or caffeic acid; these three acids are the three substrates of PAD. Thepad gene was overexpressed constitutively inEscherichia coli, and the stable purified enzyme was characterized. The difference in substrate specificity between this PAD and other PADs seems to be related to a few differences in the amino acid sequence. Therefore, this novel enzyme should facilitate identification of regions involved in catalysis and substrate specificity.

Molecular characterization and transcriptional analysis of the female-enriched chondroitin proteoglycan 2 of Toxocara canis

2017 ◽  
Vol 92 (2) ◽  
pp. 154-160
Author(s):  
G.X. Ma ◽  
R.Q. Zhou ◽  
L. Hu ◽  
Y.L. Luo ◽  
Y.F. Luo ◽  
...  
Keyword(s):  
Amino Acid ◽  
Toxocara Canis ◽  
Amino Acid Sequences ◽  
Transcriptional Analysis ◽  
Parasitic Nematodes ◽  
Biological Macromolecules ◽  
Limited Information ◽  
Reading Frame ◽  
Functional Studies ◽  
Nematode Parasites

AbstractToxocara canis is an important but neglected zoonotic parasite, and is the causative agent of human toxocariasis. Chondroitin proteoglycans are biological macromolecules, widely distributed in extracellular matrices, with a great diversity of functions in mammals. However, there is limited information regarding chondroitin proteoglycans in nematode parasites. In the present study, a female-enriched chondroitin proteoglycan 2 gene of T. canis (Tc-cpg-2) was cloned and characterized. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to measure the transcription levels of Tc-cpg-2 among tissues of male and female adult worms. A 485-amino-acid (aa) polypeptide was predicted from a continuous 1458-nuleotide open reading frame and designated as TcCPG2, which contains a 21-aa signal peptide. Conserved domain searching indicated three chitin-binding peritrophin-A (CBM_14) domains in the amino acid sequence of TcCPG2. Multiple alignment with the inferred amino acid sequences of Caenorhabditis elegans and Ascaris suum showed that CBM_14 domains were well conserved among these species. Phylogenetic analysis suggested that TcCPG2 was closely related to the sequence of chondroitin proteoglycan 2 of A. suum. Interestingly, a high level of Tc-cpg-2 was detected in female germline tissues, particularly in the oviduct, suggesting potential roles of this gene in reproduction (e.g. oogenesis and embryogenesis) of adult T. canis. The functional roles of Tc-cpg-2 in reproduction and development in this parasite and related parasitic nematodes warrant further functional studies.

Molecular characterisation and functional analysis of a cytochrome P450 gene in cotton

Biologia ◽  
2017 ◽  
Vol 72 (1) ◽  
Author(s):  
Kexue Zhou ◽  
Lu Long ◽  
Quan Sun ◽  
Weina Wang ◽  
Wei Gao ◽  
...  
Keyword(s):  
Functional Analysis ◽  
Cytochrome P450 ◽  
Verticillium Wilt ◽  
Open Reading Frame ◽  
Molecular Characterisation ◽  
Virus Induced Gene Silencing ◽  
Cytochrome P450 Gene ◽  
Reading Frame ◽  
P450 Gene ◽  
Yield And Quality

AbstractVerticillium wilt causes devastating loss of yield and quality in many crops, including cotton. To determine the molecular mechanism of resistance to verticillium wilt in cotton, we isolated a new cytochrome P450 gene, CYP94C1, and analysed its function. We obtained the complete open reading frame, which encodes a protein of 500 amino acids. The results of the functional analysis showed that resistance to verticillium wilt was enhanced when the gene was silenced using the virus-induced gene silencing (VIGS) method in cotton.

Safety and Performance of the Omnipod®Hybrid Closed-Loop System in Adolescents with Type 1 Diabetes over Five Days Under Free-Living Conditions

Diabetes ◽  
2018 ◽  
Vol 67 (Supplement 1) ◽  
pp. 1376-P
Author(s):  
GREGORY P. FORLENZA ◽  
BRUCE BUCKINGHAM ◽  
JENNIFER SHERR ◽  
THOMAS A. PEYSER ◽  
JOON BOK LEE ◽  
...  
Keyword(s):  
Type 1 Diabetes ◽  
Closed Loop ◽  
Living Conditions ◽  
Loop System ◽  
Closed Loop System ◽  
Free Living ◽  
And Performance ◽  
Free Living Conditions
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