scholarly journals Resolution and functional characterization of two mitochondrial iron-sulphur centres of the ‘high-potential iron-sulphur protein’ type

1976 ◽  
Vol 153 (1) ◽  
pp. 39-48 ◽  
Author(s):  
T Ohnishi ◽  
W J Ingledew ◽  
S Shiraishi
Keyword(s):  
Line Shape ◽  
Mitochondrial Membrane ◽  
Physiological Function ◽  
Functional Characterization ◽  
High Potential ◽  
The Other ◽  
Power Dependence ◽  
Iron Protein ◽  
Kinetic Behaviour

Two distinct iron-sulphur centres of the ‘HiPIP’ (high-potential iron-protein) type are distinguished in both pigeon heart and ox heart mitochondria. These two species, although both are paramagnetic in the oxidized state, exhibit signals which differ in their detailed line shape, field position, and temperature- and power-dependence. They also exhibit different thermodynamic and kinetic behaviour and are located on opposite sides of the mitochondrial coupling membrane. One of these centres corresponds to Centre S-3. The other ‘HiPIP’-type centre is removed readily from the mitochondrial membrane and its physiological function is not known.

scholarly journals Functional Characterization of the mazEF Toxin-Antitoxin System in the Pathogenic Bacterium Agrobacterium tumefaciens

2021 ◽  
Vol 9 (5) ◽  
pp. 1107
Author(s):  
Wonho Choi ◽  
Yoshihiro Yamaguchi ◽  
Ji-Young Park ◽  
Sang-Hyun Park ◽  
Hyeok-Won Lee ◽  
...  
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Physiological Function ◽  
Functional Characterization ◽  
Site Directed Mutagenesis ◽  
In Vitro Assays ◽  
Cell Growth Inhibition ◽  
The Right

Agrobacterium tumefaciens is a pathogen of various plants which transfers its own DNA (T-DNA) to the host plants. It is used for producing genetically modified plants with this ability. To control T-DNA transfer to the right place, toxin-antitoxin (TA) systems of A. tumefaciens were used to control the target site of transfer without any unintentional targeting. Here, we describe a toxin-antitoxin system, Atu0939 (mazE-at) and Atu0940 (mazF-at), in the chromosome of Agrobacterium tumefaciens. The toxin in the TA system has 33.3% identity and 45.5% similarity with MazF in Escherichia coli. The expression of MazF-at caused cell growth inhibition, while cells with MazF-at co-expressed with MazE-at grew normally. In vivo and in vitro assays revealed that MazF-at inhibited protein synthesis by decreasing the cellular mRNA stability. Moreover, the catalytic residue of MazF-at was determined to be the 24th glutamic acid using site-directed mutagenesis. From the results, we concluded that MazF-at is a type II toxin-antitoxin system and a ribosome-independent endoribonuclease. Here, we characterized a TA system in A. tumefaciens whose understanding might help to find its physiological function and to develop further applications.

Zebrafish tyrosylprotein sulfotransferase: molecular cloning, expression, and functional characterization

2004 ◽  
Vol 82 (2) ◽  
pp. 295-303 ◽  
Author(s):  
Emi Mishiro ◽  
Ming-Yih Liu ◽  
Yoichi Sakakibara ◽  
Masahito Suiko ◽  
Ming-Cheh Liu
Keyword(s):  
Molecular Cloning ◽  
Divalent Cations ◽  
Functional Characterization ◽  
The Other ◽  
Inhibitory Effects ◽  
Amino Acid Sequence Level ◽  
Ph Optimum ◽  
Rapid Amplification ◽  
High Degree

By employing the reverse transcriptase – polymerase chain reaction technique in conjunction with 3' rapid amplification of cDNA ends, a full-length cDNA encoding a zebrafish (Danio rerio) tyrosylprotein sulfotransferase (TPST) was cloned and sequenced. Sequence analysis revealed that this zebrafish TPST is, at the amino acid sequence level, 66% and 60% identical to the human and mouse TPST-1 and TPST-2, respectively. The recombinant form of the zebrafish TPST, expressed in COS-7 cells, exhibited a pH optimum at 5.75. Manganese appeared to exert a stimulatory effect on the zebrafish TPST. The activity of the enzyme determined in the presence of 20 mM MnCl2 was more than 2.5 times that determined in the absence of MnCl2. Of the other nine divalent metal cations tested at a 10 mM concentration, Co2+ also showed a considerable stimulatory effect, while Ca2+, Pb2+, and Cd2+ exerted some inhibitory effects. The other four divalent cations, Fe2+, Cu2+, Zn2+, and Hg2+, inhibited completely the sulfating activity of the zebrafish TPST. Using the wild-type and mutated P-selectin glycoprotein ligand-1 N-terminal peptides as substrates, the zebrafish TPST was shown to exhibit a high degree of substrate specificity for the tyrosine residue on the C-terminal side of the peptide. These results constitute a first study on the cloning, expression, and characterization of a zebrafish cytosolic TPST.Key words: zebra fish, tyrosylprotein sulfotransferase, molecular cloning.

Heterogeneity of Human Biliary Mucin: Functional Implications

1994 ◽  
Vol 86 (1) ◽  
pp. 75-82 ◽  
Author(s):  
J. Henriëtte Klinkspoor ◽  
Michel J. A. van Wijland ◽  
Carolien A. M. Koeleman ◽  
Willem van Dijk ◽  
Guido N. J. Tytgat ◽  
...  
Keyword(s):  
Cholesterol Gallstone ◽  
Lectin Binding ◽  
Gallstone Disease ◽  
Functional Characterization ◽  
The Other ◽  
Binding Studies ◽  
Human Gallbladder ◽  
Patient Heterogeneity ◽  
Model Bile

1. Human gallbladder mucin has been implicated as playing a role in the pathogenesis of gallstones. In previous studies no differences have been found in the content or composition of mucins derived from control bile or cholesterol gallstone bile. Until now, no differences were also found between these two groups of mucins with regard to their ability to cause cholesterol nucleation. In the accompanying paper we have reported that there is a strong heterogeneity of gallbladder mucins derived from individual patients (M. J. A. van Wijland, J. H. Klinkspoor, L. Th. de Wit, R. P. J. Oude Elferink, G. N. J. Tytgat and A. K. Groen, Clin Sci 1994; 86: 67–74). In the present study we further investigated a possible patient to patient heterogeneity of mucin by means of immunological and functional characterization of mucins isolated from hepatic bile of six different patients with gallstones. 2. Considerable heterogeneity was found. Two of the mucins barely reacted with a polyclonal anti-mucin antibody, whereas the other four mucins reacted very strongly. Lectin-binding studies indicated that the glycans of these two mucins expressed less D-N-acetylgalactosamine residues than the other four mucins. This was confirmed by analyses of the glycan compositions. These studies furthermore indicated that the glycans were of the O-linked type, contained α-D-N-acetylglucosamine and were fucosylated, sialy-lated and sulphated to different extents. Except for a strong heterogeneity in the sugar composition of the mucins, heterogeneity was also found in the biological activity of the mucins. The two immunologically diverging mucins nucleated cholesterol from model bile 1–2 days earlier and also caused an almost threefold more rapid rupture of cholesterol/phospholipid vesicles than the other mucins. 3. We conclude that considerable differences exist between mucins derived from individual patients and that the heterogeneity in glycan composition might play a role in the pathogenesis of gallstone disease.

scholarly journals Characterization of βαβββ Resistance Proteins from Pseudomonas aeruginosa

2014 ◽  
Vol 70 (a1) ◽  
pp. C717-C717
Author(s):  
Allegra Vit ◽  
Monika Popp ◽  
Eyad Kalawy-Fansa ◽  
Shen Yu ◽  
Wulf Blankenfeldt
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Physiological Function ◽  
Functional Characterization ◽  
Conserved Residues ◽  
X Ray ◽  
Resistance Proteins ◽  
X Ray Crystallography ◽  
Phenazine Derivative ◽  
Function Assignment

Pseudomonas aeruginosa is a multiresistant pathogen that can cause infection in immuno-compromized patients, for example in people suffering from cystic fibrosis. [1] It has complex patho-physiology and produces a large number of exoproducts, among which the phenazines are especially prominent. In P. aeruginosa, the blue phenazine derivative pyocyanin plays a crucial role in infection of the host. [2] This phenazine can generate reactive oxygen species and is thought to act as respiratory pigment and as a virulence factor at the same time. P. aeruginosa has to protect itself from its own phenazines because of the antibiotic action of these substances. Inspired by the fact that the phenazine biosynthesis operon of several bacteria contains a phenazine resistance factor of the βαβββ module protein family, we have searched the genome of P. aeruginosa for proteins of this fold. [3] In P. aeruginosa we could identify 22 of these genes, most without previous functional characterization. A structure-based sequence alignment made it possible to assign these proteins to two classes with two subgroups each, based on the conserved residues in the active site. Using X-ray crystallography and biophysical methods, we further demonstrate that several of these proteins indeed bind phenazines and possibly other antibiotics that contain aromatic moieties. Currently, we are working on the structural characterization and physiological function assignment of all of these βαβββ-module-containing proteins. Ultimately, these data may lead to novel anti-infective strategies.

scholarly journals Heterogeneous clinical features in Cockayne syndrome-A patients with the same mutation and in siblings

2021 ◽  
Author(s):  
Asma CHIKHAOUI ◽  
Ichraf Kraoua ◽  
Nadège Calmels ◽  
Sami Bouchoucha ◽  
Cathy Obringer ◽  
...  
Keyword(s):  
North Africa ◽  
Clinical Symptoms ◽  
Excision Repair ◽  
Functional Characterization ◽  
Clinical Manifestations ◽  
Case Series ◽  
Cockayne Syndrome ◽  
The Other ◽  
Indel Mutation

Abstract Background Cockayne syndrome (CS) is a rare autosomal recessive disorder caused by mutations in ERCC6/CSB or ERCC8/CSA that participate in transcription-coupled nucleotide excision repair (TC-NER) of UV-induced DNA damage. CS patients display a large heterogeneity of clinical symptoms and severities, the reason of which is not fully understood, and little data is available for affected siblings. CS is largely undiagnosed in North Africa. Methods We report here the clinical description as well as genetic and functional characterization of eight North African CS patients, including siblings. These patients, who belonged to six unrelated families, underwent complete clinical examination and biochemical analyses. Sanger sequencing was performed for the recurrent mutation in five families, and targeted gene sequencing for one patient of the other family. We also performed RRS (Recovery RNA Synthesis) to confirm the functional impairment of DNA repair in the identified mutations. Results Six out of eight patients carried a homozygous indel mutation (c.598_600delinsAA) in exon 7 of ERCC8, and displayed a variable clinical spectrum, including between siblings, despite sharing the same mutation. The other two patients were Tunisian siblings who carried a homozygous splice-site variant in ERCC8 (c.843 + 1 G > C). They presented more severe clinical manifestations, which are in general rarely associated with CSA mutations, leading to gastrostomy and hepatic damage. Impaired TC-NER was confirmed by RRS in six tested patients. Conclusions This study provides the first deep characterization of case series of rare CS-A patients in North Africa. They carry mutations described to date only in this region and the Middle-East. We also provide the largest characterization of unrelated patients, as well as siblings, with the same mutation, providing a framework for dissecting elusive genotype-phenotype correlations in CS.

scholarly journals Characterization of the glucose 6-phosphate dehydrogenase activity in rat liver mitochondria

1976 ◽  
Vol 159 (3) ◽  
pp. 683-687 ◽  
Author(s):  
M Grunwald ◽  
H Z Hill
Keyword(s):  
Dehydrogenase Activity ◽  
Rat Liver ◽  
Liver Mitochondria ◽  
The Other ◽  
Rat Liver Mitochondria ◽  
Kinetic Behaviour ◽  
Two Peaks ◽  
Glucose 6 Phosphate Dehydrogenase

Glucose 6-phosphate dehydrogenase activity in rat liver mitochondria can be released by detergent. The released activity is separated by chromatography into two peaks. One peak has the kinetic behaviour and mobility similar to the soluble sex-linked enzyme, whereas the other peak is similar to the microsomal hexose 6-phosphate dehydrogenase. There is no evidence for the existence of a new glucose 6-phosphate dehydrogenase activity in rat liver mitochondria.

scholarly journals FUNCTIONAL PROPERTIES OF TABLE SUGARS DERIVED FROM THE SAP OF THE INFLORESCENCES OF 03 COCONUT (COCOS NUCIFERA.L) CULTIVARS IN COTE DIVOIRE

2020 ◽  
Vol 8 (11) ◽  
pp. 254-263
Author(s):  
Okoma D. Muriel J. ◽  
◽  
Konan K. Jean Louis ◽  
Assa Rebecca R ◽  
◽  
...  
Keyword(s):  
Functional Characterization ◽  
Cane Sugar ◽  
Polyphenolic Compounds ◽  
The Other ◽  
Total Polyphenols ◽  
Coconut Tree ◽  
White Cane ◽  
Crystalline Sugars ◽  
Cooking Times

This study is part of a context of diversification of the uses of the Ivorian coconut tree. The objective was to determine the functional characteristics of crystalline sugars derived from the sap of inflorescences of three coconut cultivars. Red and white sugars from cane were taken as controls. Variations in final temperatures coupled with distinct cooking times were carried out in order to evaluate the effect of the time/temperature couple on the studied parameters. Thus, three different treatments were applied.The functional characterization of the sugars studied shows that coconut sugars are an important source of total polyphenols with levels ranging from 34.64 to 143.12 mg/100g.No polyphenolic compounds were assayed in white cane sugar. Coconut sugars from treatment 1 are less energetic than those from the other two treatments. On the other hand, brown and white sugars from sugar cane are more energetic than those from coconut trees.In view of all the above, coconut sugars, especially those from treatment 1, are natural sweeteners with a low energy value. In addition, they are rich in polyphenols and flavonoids, unlike refined cane sugar and its red counterpart which contains very few nutrients.Thus, coconut sugars produced in Cote dIvoire can be considered as a phytonutrient substitute, capable of replacing sugarcane sugars.

scholarly journals Functional characterization of SARS-CoV-2 infection suggests a complex inflammatory response and metabolic alterations

2020 ◽  
Author(s):  
Lucía Trilla-Fuertes ◽  
Ricardo Ramos ◽  
Natalia Blanca-López ◽  
Elena López-Camacho ◽  
Laura Martín-Pedraza ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Functional Characterization ◽  
Self Regulation ◽  
Transcriptional Response ◽  
The Other ◽  
Differentially Expressed ◽  
Node Activity ◽  
Cytokines And Chemokines ◽  
Variable Genes

AbstractCovid-19, caused by the SARS-CoV-2 virus, has reached the category of a worldwide pandemic. Even though intensive efforts, no effective treatments or a vaccine are available. Molecular characterization of the transcriptional response in Covid-19 patients could be helpful to identify therapeutic targets. In this study, RNAseq data from peripheral blood mononuclear cell samples from Covid-19 patients and healthy controls was analyzed from a functional point of view using probabilistic graphical models. Two networks were built: one based on genes differentially expressed between healthy and infected individuals and another one based on the 2,000 most variable genes in terms of expression in order to make a functional characterization. In the network based on differentially expressed genes, two inflammatory response nodes with different tendencies were identified, one related to cytokines and chemokines, and another one related to bacterial infections. In addition, differences in metabolism, which were studied in depth using Flux Balance Analysis, were identified. SARS-CoV2-infection caused alterations in glutamate, methionine and cysteine, and tetrahydrobiopterin metabolism. In the network based on 2,000 most variable genes, also two inflammatory nodes with different tendencies between healthy individuals and patients were identified. Similar to the other network, one was related to cytokines and chemokines. However, the other one, lower in Covid-19 patients, was related to allergic processes and self-regulation of the immune response. Also, we identified a decrease in T cell node activity and an increase in cell division node activity. In the current absence of treatments for these patients, functional characterization of the transcriptional response to SARS-CoV-2 infection could be helpful to define targetable processes. Therefore, these results may be relevant to propose new treatments.

scholarly journals Identification of Loci and Functional Characterization of Trichothecene Biosynthesis Genes in Filamentous Fungi of the Genus Trichoderma

2011 ◽  
Vol 77 (14) ◽  
pp. 4867-4877 ◽  
Author(s):  
R. E. Cardoza ◽  
M. G. Malmierca ◽  
M. R. Hermosa ◽  
N. J. Alexander ◽  
S. P. McCormick ◽  
...  
Keyword(s):  
Functional Analysis ◽  
Gene Rearrangement ◽  
Gene Loss ◽  
Functional Characterization ◽  
The Other ◽  
Regulatory Proteins ◽  
Biosynthetic Gene ◽  
Content Type ◽  
Cluster Organization

ABSTRACTTrichothecenes are mycotoxins produced byTrichoderma,Fusarium, and at least four other genera in the fungal orderHypocreales.Fusariumhas a trichothecene biosynthetic gene (TRI) cluster that encodes transport and regulatory proteins as well as most enzymes required for the formation of the mycotoxins. However, little is known about trichothecene biosynthesis in the other genera. Here, we identify and characterizeTRIgene orthologues (tri) inTrichoderma arundinaceumandTrichoderma brevicompactum. Our results indicate that bothTrichodermaspecies have atricluster that consists of orthologues of seven genes present in theFusariumTRIcluster. Organization of genes in the cluster is the same in the twoTrichodermaspecies but differs from the organization inFusarium. Sequence and functional analysis revealed that the gene (tri5) responsible for the first committed step in trichothecene biosynthesis is located outside the cluster in bothTrichodermaspecies rather than inside the cluster as it is inFusarium. Heterologous expression analysis revealed that twoT. arundinaceumcluster genes (tri4andtri11) differ in function from theirFusariumorthologues. TheTatri4-encoded enzyme catalyzes only three of the four oxygenation reactions catalyzed by the orthologous enzyme inFusarium. TheTatri11-encoded enzyme catalyzes a completely different reaction (trichothecene C-4 hydroxylation) than theFusariumorthologue (trichothecene C-15 hydroxylation). The results of this study indicate that although some characteristics of thetri/TRIcluster have been conserved during evolution ofTrichodermaandFusarium, the cluster has undergone marked changes, including gene loss and/or gain, gene rearrangement, and divergence of gene function.

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