bluuebunny/crossref_metadata_2025 huggingface dataset:
In silico Identification and Analysis of Iron (Fe) Transporters in Various Plant Species
Authors: Dinesh Akula , Rathod Ramya , Sreedhar M.
Published: December 2020

In plants, Iron is an important micronutrient which is required for various processes like photosynthesis, respiration and for balanced redox potential. Iron has a significant role in human nutrition. Therefore, increasing the Fe content of economical parts through conventional, Molecular and/or Transgenic breeding will have dramatic impact on human health. Therefore, this study was aimed to identify and characterize homologues of OsIRT1, OsIRT2, OsVIT1, OsYSL2, OsYSL15 and OsYSL18 in 21 different plant species. The study revealed that, a total of 51 putative Fe transporter proteins homologues were identified which could be characterized with 555 to770 amino acids sequence length with 61 to 84.8 kDa molecular weight and pI of 8.2 to 9.4 having basic nature with 9 to15 TMD with an average of 13 TMDs. The sub-cellular localization of putative Fe transporters was predicted as the plasma membrane and all the identified Fe homologues contained oligopeptide transporter (OPT) domain proteins which play a significant to role in Fe transport and homeostasis. Further, the analysis led to identification of highly conserved residues in the putative Fe sequences which could be used as potential motif signatures in identification of new Fe transporter. The interactome analysis for oryza sativa OsYSL15 transporter showed putative interaction with NAS2, NAS1, DMAS1, OsJ_32857, IRT1, IRO2, IDEF1, IRT2, IDI2 which are found to be directly involved in Fe transport from roots to grain. This study, elucidates the valuable theoretical knowledge about the Fe genes, protein features and assist in molecular manipulation of Fe transporters in various plants for developing high Fe in economical part.
Livelihood Dependence on Highland Pastoralism (Doksa) in Trans-Himalayan Region of Zanskar, Ladakh
Authors: Raj Anup , Angchuk Dorjey , Islam M. A.
Published: December 2020

A unique system of short-distance vertical transhumance pastoralism has evolved in the Trans-Himalayan region of Zanskar, Ladakh in response to short summer cropping season and vast alpine pasturelands. Cattle are taken to highland pastures for three-and-a-half month in summer season and kept in temporary settlements locally called as doksa. The study investigated the herding practices, migration pattern, livestock production and livelihood dependence on highland pastoralism. Purposive sampling technique was administered to withdraw the sample of 6 doksas and the data were collected from both secondary and primary sources. Results revealed that 31 herders in the 6 doksas possessed a total of 794 milk producing zhomos. The doksas produced 3 lakh litres of milk, 18000 kg of butter and the same amount of dried protein cake (chhurpey). The herders earned incomes of ₹ 122500.00 and ₹ 59375.00 by trading 300-350 kg of butter and 450-500 kg of chhurpey, respectively with employment opportunities of 3100 woman-days/year. The poor living conditions at doksa and unavailability of alternative economic opportunities for women herders has threatened its continuation for cash generation, food and livelihood security and socioeconomic development. Therefore, livelihood diversification using alternative resources is imperative to keep pace with current development and future challenges.
Relationship of Clinical Data and Confirmed Case of Disease by the New Coronavirus, in the Mexican State of Guanajuato
Authors: Raygoza Nicolás Padilla- , Vargas Gilberto Flores- , Olivos Efraín Navarro- , Lona Elia Lara- , Luna María de Jesús Gallardo- , Vázquez Francisco Javier Magos- , Martínez Daniel Alberto Díaz-
Published: December 2020

Aims: The objective was to analyze the clinical data in population from Mexican Guanajuato state as a suspected case of COVID-19 and with result positive of rRT-PCR, reported until October 2, 2020.

Study Design: It is a cross-sectional based in data from National Epidemiological Surveillance System from General Epidemiological Directorate, Secretary of Health in Mexico.

Place and Duration of Study: Sample: All registries from confirmed and discarded cases of COVID-19 in database until October 2, 2020.

Methodology: It was included 100,919 registries, and from them, 810 were excluded by missing the result of rRT-PCR test. A suspected case was one with a clinical finding considered greater (cough, fever, headache or dyspnea and accompanied by at least one of the following: myalgia, arthralgia, odynophagia, chills, chest pain, rhinorrhea, anosmia, dysgeusia or conjunctivitis); a confirmed case of COVID-19 is a person with a positive rRT-PCR test for SARS-CoV-2, regardless of the clinical data presented. We included age, sex, and clinical data registered and result of rRT-PCR for SARS-CoV-2. It was used logistic regression to analyze the effect of clinical data on positive rRT-PCR.

Results: It was analyzed 100,109 registries. From them, 41,734 were positive for SARS-CoV-2. Fever (OR 1.72, CI95% 1.68 to 1.77), cough (OR 1.70, CI95% 1.66 to 1.74), and odynophagia (OR 1.71, CI95% 1.66 to 1.75) shown a stronger effect on positive rRT-PCR test. Cyanosis did not have any effect on the result of the rRT-PCR test.

Conclusion: There are not pathognomonic clinical data for COVID-19. All clinical data in confirmed cases are similar to another respiratory viral infections.
Psychiatric Comorbidities among Children with Attention Deficit Hyperactivity Disorders
Authors: Shaaban Sahar Sayed Ahmed , Seleem Mohammad Abd El-Hakeem , Badawy Adel Abd El –Kareem , Eissa Mai Abd El–Raouf
Published: December 2020

Background and Purpose: Attention Deficit Hyperactivity Disorder (ADHD) is the most common neurodevelopmental disorder in children. The purpose of this study is to explore the rate of psychiatric comorbidities among children with ADHD and investigating the impact of comorbidities on function of children.

Subjects and Methods: Sixty-four children of both genders who fulfilled DSM-5 criteria for ADHD were included. All subjects were subjected to IQ assessment, Kiddie Schedule for Affective Disorders and Schizophrenia (K-SADS), Conners Rating Scale, and Child Behavior Check List (CBCL).

Results: Combined subtype was the commonest (50%) followed by hyperactive-impulsive (31.25%), then inattentive (18.75%). Psychiatric comorbid disorder among pediatric cases with ADHD was ODD (31.25%), followed by nocturnal enuresis (23.44%), conduct disorders (18.75%), motor tic disorders (15.63%), anxiety disorders (12.50%), depressive disorders (10.94%), substance abuse disorders (9.38%), autism spectrum disorders (7.81%), vocal tic disorders (1.56%).  There was a statistically significant difference between gender and conduct disorder and SUDs (P <0.05). There was no significant difference in the distribution of comorbidities among subtypes except for conduct disorder (P<0.05). There was a significant difference between comorbidities and severity of ADHD, according to the Conners scale (P<0.05). There was a significant difference between comorbidities number and different CBCL parameters (P<0.05). There was a significant negative correlation between the number of comorbidities and total competence and positive correlation with the total problem (P<0.05).

Conclusions: The presence of comorbid disorders among pediatric cases with ADHD is the rule rather than the exception. Oppositional defiant disorder, followed by nocturnal enuresis, are the most common comorbid diseases. Comorbidities number is negatively correlated with total competence and positively correlated with the total problem.
Antimicrobial Activity of Antibiotic Doped Carbon Nanoparticles Extracted from Kitchen Soot against Pathogenic Gram-negative Bacteria
Authors: Habiba . , Din Israf Ud , Tasleem S. , Mubashir Muhammad , Bakht M. Afroz
Published: December 2020

In this study carbon nanoparticles were extracted from kitchen soot and characterized by using UV/Visible spectroscopy.  Amoxicyline and vibramycin were doped on the carbon nanoparticles by mixing the solutions of the aforementioned antibiotics and carbon nanoparticles and then evaporated to dryness at 50°C. The antibacterial potential of carbon nanoparticle, antibiotics and carbon nanoparticles doped antibiotics were evaluated using Agar tub dilution method against two bacterial strain i.e. P. Aerogunis and Salmonella entrica. The result indicated that the carbon nanoparticle doped antibiotics showed marked increase in the antibacterial potential with inhibition zone of 16.2 mm against P. aerogunisa and 12.5 mm against Salmonela entrica for Vibramycin Doped CNPs. The inhibition zone of Amoxicyline doped carbon nanoparticle is 25.0 mm against P. aerogunisa and 30.0 mm against Salmonela entrica.
Effectiveness of Teaching by SWOM Strategy in Achievement and Retention of Second Intermediate-Grade Students in Science
Authors: Hussein Hanan Nassir , Mater Firas Ayal
Published: December 2020

This research-aims to investigate (the effectiveness of teaching SWOM strategy in achievement and second-graders intermediate students retention in the sciences subject) The experimental design (with-partial-control) was chosen for two equivalent groups, experimental and control, the sample was randomly selected-from-students (Hittin intermediate school( for girls in Thi-Qar Governorate), the groups were equivalent in-variables (Age calculated in months – female students’ achievement in-science for the first-grade (previous-year) –previous information testin science- subject – intelligence test (Raven), behavioral objectives in cognitive domain were set reached (100) behavioral objectives, the necessary teaching plans were prepared for two research. The research tools, the researcher prepared a test comprises (40) MC items with four alternatives. Validity, reliability, discrimination-coefficient, difficulty-and-alternatives effectiveness were calculated. At the end of second semester (2018-2019),the experiment was applied. After the experiment was completed, the achievement test was applied on two research groups. Then, the same achievement test was applied again after a two weeks period-from first application to check information retention, and for processing statistical data, the statistical methods were used (t-test for two independent samples, difficulty-equations, items-discrimination, wrong alternatives-effectiveness and Pearson-correlation-equation), the-research was reached the results: There is a statistical significant difference at the significance level (0.05) between second grade students means scores who study science subject according-to the SWOM strategy and the second-grade students mean-scores who study the-same subject by traditional method in achievement and retention test in favor of the experimental group, depend on the results, several conclusions were formulated, conclusions and recommendations-were reached, to complete th- research, several studies-were proposed.
Effect of Using Scientific Stations Strategy in Developing Deductive Thinking of Intermediate School Students in General Sciences
Authors: Al-Hafidh Heba Mohammed Saleh
Published: December 2020

The study aimed to know the effect of using the Scientific stations strategy in developing Deductive thinking of intermediate school students in the science subject. To achieve the aim of the research, the researcher used experimental design with two equal groups, the research sample consisted of (65) intermediate-grade students in Al Nidhal school for boys represented by (30) students of the experimental group and (35) students for the control group, the researcher equivalence of the two research groups with a set of variables including (Intelligence, parents ’educational level, previous achievement, age calculated by months and pre-test for Deductive thinking), to verify the research aim the researcher put the two hypotheses as following : 1- There is no statistical significant difference at the level of significance (0.05) between the scores mean of the members of the experimental group and the members of the control group in the post test of Deductive thinking. 2- There is no statistical significant difference at the level of significance (0.05) between the mean difference between the (pre and post) tests of both experimental and control groups in the Deductive thinking test. To verify the research hypotheses, the researcher prepared a test for Deductive thinking consisting of (20) items the researcher applied the test of Deductive thinking, collected the data, and tabulated it in tables for statistical processing and the results indicated: 1- There is a statistical significant difference between the scores mean of students in both groups in the Deductive thinking test and in favor of the experimental group. 2- There is a statistical significant difference between the mean difference between the two tests (pre - post) and in favor of the experimental group.
Martin Luther King Jr’s Delivery Style in Political Context: A Critical Discourse Analysis
Authors: Naser Mutar Sabeeh , Katea Dr. Haider Hussein , Hammadi Majid Dakhil
Published: December 2020

The effect(s) of delivery style may be in the form of punctuation and accentuation of speech, or they may be of subtler nature like adding emotion and force to utterance(s). This article then, investigates the position of paralinguistic factors: pace of delivery, pitch, intensity and pause pattern in King Martin Luther’s speech “Have a Dream” in political context. These factors carry attitude, psychological condition and emotional state of the speaker to the audience and affect their attitudes and emotions. These factors have been called the factors of delivery style because they can be used only in spoken language. These factors are the physical properties of speech and thus they are observable. These factors, thenwill be analyzed and studied with the help of van Dijk Model of CDA to answer To what extent the ideology of Martin Luther King Jr is reflected through the paralinguistic choices “I Have Dream”. In terms of the findings, the analysis found that persuasion is not the result of the use of the persuasive strategies in words and syntax alone as delivery style also plays an important role in persuasion. Pace of delivery along with pitch intensity and pause pattern are the main components of political speech delivery style for persuasive end. Moreover, the delivery style adds certain effect to other rhetorical strategies. Martin Luther King Jr. is a master of speech delivery style as he makes the best persuasive use of pace, pause, pitch and intensity in his speech.
The Adaption of Self-Reflexivity and Metafiction Approach to Myth and History in Shashi Tharoor's the Great Indian Novel: A Post-Modernist Study
Authors: Rahman Suzan Raheem , Rasheed Lamiaa Ahmed , Mustafa Lujain Ismael
Published: December 2020

Shashi Tharoor's The Great Indian Novel is an example of a post-modern historiographic metafiction that takes the relationship between reality and fiction into consideration. This novel also depicts the 20th century political past by reviving events, incidents and characters of the myth of Mahabharata. The current paper aims to explain how Tharoor rebuilds the twentieth-century past by drawing on the great Mahabharata classical epic. Additionally, it examines the common relationship between fiction and history as it progressed along and continuous processes through the use of self-reflexivity and metafiction approach. In The Great Indian Novel, Tharoor adapts a metafiction tool which is the most fitting way to tackle this novel as a postmodernist study. Tharoor blends fiction and fact through a self-reflective narrative and the use of several metafiction devices by adapting the myth of Mahabharata to construct the distance between the past and the present. Tharoor takes the ancient myth as the basic structure with contemporary group of political characters for a real and ironic review of recent Indian history and representation.
Using Video Modeling and Video Prompting to Teach Conversational Skills to Students with Autism: A Consideration of Effectiveness, Practicality, and Acceptability
Authors: Almalki Saeed
Published: December 2020

Many studies have been focused on video modeling and prompting as effective evidence-based practices to teach conversational skills to students with autism spectrum disorder (ASD). However, few studies have focused on teachers who implement video modeling and prompting in their classrooms. This article aims to describe the experiences and perceptions of four special education teachers who have used video modeling and prompting with four autistic students for a semester in their classrooms. The participants were asked questions related to three main areas: effectiveness, practicality, and acceptability. Most of the teachers did not regard this intervention as an effective and acceptable way to teach conversational skills to students with ASD. Training and support was requested by all teachers to enhance the practicality of this intervention. Other important implications for researchers and teachers were identified.
The Effects of Positive Parenting toward Intolerance in Pre-School Children
Authors: Latipah Eva , Kistoro Hanif Cahyo Adi , Putranta Himawan
Published: December 2020

The absence of mutual respect and tolerance in preschoolers is getting stronger. This research aims to elaborate on the manifestation and causes of intolerance and examine the effect of positive parenting on intolerance. This research is field research using survey and experimental methods. The survey was been conducted to identify teacher needs as a basis for developing a positive parenting module. Experiments were been conducted to see the effect of positive parenting on intolerance. The subjects of this research were 129 pre-school children in Yogyakarta who were been selected using a purposive sampling technique. Data analysis uses qualitative analysis techniques (reduction, display, and conclusion) and quantitative (t-test). The results showed Intolerance of many pre-school children occurs around religion and morals (blaming and hating the religion of different friends, not wanting to share and make friends with friends who are different from themselves). Causes of intolerance include imitating songs, watching television, and the internet, imitating parents, friends, or teachers. Positive parenting affects the intolerance of pre-school children, as indicated by the value of t = 3.396 and p = 0.000, which means that positive parenting can reduce pre-school children's intolerance. Therefore, sustainable positive parenting needs to be implemented within both the family, school, and community, so that intolerance in pre-school children does not arise.
Inclusive Education Practices for Refugee Children with Disabilities in Jordanian Schools
Authors: Muhaidat Mohammad , Alodat Ali M. , Almeqdad Qais I.
Published: December 2020

Jordanian schools have hosted thousands of Syrian refugees in the last decade. This qualitative study aimed to identify inclusive education practices used with Syrian refugee children with disabilities in Jordanian schools. Ten schools were evaluated using observation and data analysis methods and semi-structured interviews with four teachers and five refugee children with disabilities' parents. This study's results revealed that Jordanian schools lack the minimum inclusive education standards for refugee children with disabilities regarding legislation, accessibility, modifications, and accommodations. Four themes have emerged from the data, observations, and interviews' analysis that represent the main challenges facing the inclusive education practices in refugee children with disabilities schools. These themes were: schools' capacity, financial limitations, program availability, and teachers' qualifications. Finally, a set of recommendations and suggestions to enhance the best practices of inclusive education for refugee children with disabilities were provided.
Possible physiological role of epidermal growth factor in the development of the mouse mammary gland during pregnancy
Authors: Taketani Yuji , Oka Takami
Published: July 2002

Epidermal growth factor stimulated cell proliferation in a primary mammary epithelial cell culture derived from mice at different stages of pregnancy. Moreover, the peptide hormone inhibited casein production induced by the synergistic actions of insulin, cortisol and prolactin. The inhibitory effect of epidermal growth factor was influenced by the gestational stages of the mammary gland. These effects of epidermal growth factor were exerted at physiological concentrations. The dual actions of epidermal growth factor on mammary cells implicate its participation in regulation of the growth and differentiation of the mammary gland during pregnancy.
Stimulation of specific GTPase activity by vasopressin in isolated membranes from cultured rat hepatocytes
Authors: Fain John N. , Brindley David N. , Pittner Richard A. , Hawthorne John N.
Published: July 2002

Membranes were isolated by isotonic homogenization and differential centrifugation from rat hepatocytes cultured overnight. The specific GTPase activity of the membranes was 1–1.3 pmol γ‐labelled GTP hydrolysed/mg protein per min in the presence of 1.2 mM Na+, 2 mM EGTA, 1 mM ATP and 0.2 mM 5‐adenylyl imidodiphosphate. Under these conditions there was a stimulation of specific GTPase activity of no more than 20% by 11–115 nM vasopressin. No effect of vasopressin was seen in the presence of 1.7 μM free Ca2+ or 100 mM Na+. The findings indicate that vasopressin is able to influence GTPase activity as well as accelerate phosphoinositide breakdown in rat hepatocytes.
Regulation of a liver plasma membrane phosphoinositide phosphodiesterase by guanine nucleotides and calcium
Authors: Uhing Ronald J. , Jiang Hang , Prpic Veronica , Exton John H.
Published: July 2002

Rat liver plasma membranes are enriched in a Ca2+‐dependent phosphodiesterase active on phosphatidylinositol 4,5‐P2 and phosphatidylinositol 4‐P, but not phosphatidylinositol. Inositol‐P3 is the first product of the reaction, but is rapidly degraded. Micromolar concentrations of GTP and its nonhydrolyzable analogues stimulate the reaction, whereas GDP, GMP and other nucleoside triphosphates are inactive. GTP and its analogues decrease the requirement of the reaction for Ca2+ and also increase its activity at saturating Ca2+. These results support the hypothesis that guanine nucleotides and a guanine nucleotide binding regulatory protein are involved in coupling the receptors for Ca2+‐mediated agonists to the breakdown of plasma membrane phosphatidylinositol 4,5‐P2.
Neomycin: a specific drug to study the inositol‐phospholipid signalling system?
Authors: Prentki Marc , Deeney Jude T. , Matschinsky Franz M. , Joseph Suresh K.
Published: July 2002

Neomycin, an antibiotic previously thought to interact specifically with inositol‐containing phospholipids, was found to inhibit IP3‐mediated Ca2+ release from the intracellular stores of permeabilized insulinoma and liver cells. This inhibition could be relieved by increasing the IP3 concentration. Radiolabelled IP3 was found to bind tightly to columns prepared from neomycin covalently attached to glass beads. ATP was also bound by these colums. It is concluded that neomycin acts in biological systems as a weak anion exchanger and is therefore unsuitable for use as a specific tool to study the role of inositol phospholipids in intracellular signalling.
Adjacent codon‐anticodon interactions of both tRNAs present at the ribosomal A and P or P and E sites
Authors: Rheinberger Hans-Jörg , Nierhaus Knud H.
Published: July 2002

A labeled tRNA present at the A, P or E site can be partially chased from the ribosome, a cognate non‐labeled tRNA as chasing substrate being 3‐12‐times more efficient than non‐cognate tRNA at a molar ratio tRNA: 70 S = 10:1. These findings indicate that a tRNA bound to a programmed ribosome undergoes codon‐anticodon interaction at all three sites (A, P and E site). Furthermore, both labeled tRNAs present on the ribosome can be chased more effectively with cognate than with non‐cognate substrate at the same time. This finding provides strong evidence that both tRNAs present on the ribosome exhibit simultaneous codon‐anticodon interaction. This is valid for both the pretranslocational state (Ac[3H]Lys‐tRNALys in the A and [14C]tRNALys in the P site) as well as the posttranslocational state (Ac[3H]Lys‐tRNALys in the P and [14C]tRnaLys in the E site).
Presence of tyrosine‐O‐sulfate in sheep pituitary prolactin
Authors: Kohli Rita , Chadha Neeraja , Muralidhar K.
Published: July 2002

When the metabolically obtained 35S‐labelled sheep pituitary prolactin‐rich fraction was subjected to chemical deglycosylation the radioactivity was retained in the immunoprecipitable prolactin. 35S‐labelled prolactin‐rich pituitary extract was fractionated on SDS‐PAGE and protein was extracted from prolactin positive bands. When the extracted 35S‐labelled prolactin was hydrolysed by alkali and then chromatographed on a thin layer of silica, it showed the presence of a radioactive compound which had an R
f value identical to the standard Tyr‐O‐SO4 synthesized and characterized in our laboratory.
Atrial natriuretic peptide (ANP) in rat gastrointestinal tract
Authors: Vuolteenaho O. , Arjamaa O. , Vakkuri O. , Maksniemi T. , Nikkilä L. , Kangas J. , Puurunen J. , Ruskoaho H. , Leppäluoto J.
Published: July 2002

Acid extracts of rat stomach and small intestine contained 8.6 ± 3.7 and 39 ± 15 ng/g of immunoreactive atrial natriuretic peptide (ANP). When studied by gel filtration and reverse‐phase high‐performance liquid chromatography, the stomach immunoreactivity consisted of multiple components, whereas the small intestine contained mostly proANP and ANP 1–28‐like material. These findings indicate that ANP may have a role in the physiology of the gastrointestinal tract, e.g. in the regulation of water and electrolyte absorption.
Mitochondrial oxidative phosphorylation participating in photosynthetic metabolism of a leaf cell
Authors: Krömer Silke , Stitt Mark , Heldt Hans W.
Published: July 2002

Oligomycin was used for the selective inhibition of mitochondrial oxidative phosphorylation in barley leaf protoplasts. The addition of oligomycin inhibited the photosynthesis of intact protoplasts by 40–60%, this being accompanied by a pronounced decrease in cellular ATP content. When the protoplasts were ruptured in a way leaving the chloroplasts intact, the photosynthesis of the released chloroplasts was not inhibited by oligomycin. It is therefore concluded that in illuminated protoplasts, ATP formation by oxidative phosphorylation is required for utilization of the photosynthate by the cell. Thus, mitochondrial oxidative phosphorylation appears to serve an essential function for supplying the cytosol with ATP during photosynthesis.
Purification of the human placental α2‐macroglobulin receptor
Authors: Jensen P.H. , Moestrup S.K. , Gliemann J.
Published: July 2002

The α2‐macroglobulin receptor was solubilized from human placental membranes, purified and characterized. Affinity cross‐linking of labelled ligand to intact membranes showed a receptor size compatible with 400–500 kDa. The membranes were solubilized in 3‐[(3‐cholamidopropyl)dimethylammonio]propane sulfonate (CHAPS) and affinity chromatography was performed using Sepharose‐immobilized α2‐macroglobulin‐methylamine with elution in buffer containing 2 mM EDTA, pH 6.0. SDS‐PAGE of the resulting receptor preparation showed a predominant approx. 440 kDa band (reducing conditions) and some minor accompanying proteins of 70–90 kDa and 40 kDa. The yield was 400–800,μg receptor preparation per placenta. The receptor preparation immobilized on nitrocellulose bound the α2‐macroglobulin‐trypsin complex with a dissociation constant of about 400 pM. 125I‐iodinated receptor preparation bound almost quantitatively to Sepharose‐immobilized α2‐macroglobulin‐methylamine in the presence of CHAPS alone, and bound 70–80% in the presence of 0.2% SDS. The labelled proteins were separated in the presence of 0.2% SDS by gel filtration or SDS‐PAGE (unboiled samples). The 440 kDa protein accounted for the major part of the binding, although some approx. 80 kDa proteins, perhaps proteolytic degradation products, also showed binding activity.
The interaction of actin with dystrophin
Authors: Levine B.A. , Moir A.J.G. , Patchell V.B. , Perry S.V.
Published: July 2002

Proton NMR spectroscopy of synthetic peptides corresponding to defined regions of human dystrophin has been employed to study the interaction with F‐actin. No evidence of interaction with a C‐terminal region corresponding to amino acid residues 3429–3440 was obtained. F‐actin restricted the mobility of residues 19–27 in a synthetic peptide corresponding to residues 10–32. This suggests that this is a site of F‐actin interaction in the intact dystrophin molecule. Identical sequences to that of residues 19—22 in dystrophin, namely Lys‐Thr‐Phe‐Thr are also present in the N‐terminal regions of the α‐actinins implying this is also a site of F‐actin interaction with α‐actinin.
Secretion of apolipoprotein B in serum‐free cultures of human hepatoma cell line, HepG2
Authors: Adeli Khosrow , Sinkevitch Cheryl
Published: July 2002

We have developed a defined medium which can maintain efficient growth of HepG2 cells sustaining the synthesis of a variety of plasma proteins including apolipoprotein B. This defined system was used to investigate long‐term effects of insulin, estrogen, triiodothyronine, cholesterol, and oleate on the growth pattern of HepG2 cells and secretion rate of apolipoprotein B. Oleate and triiodothyronine caused significant increases in secretion of apolipoprotein B. The stimulatory effect of triiodothyronine was only observed after long (6 days) exposure of cells to the hormone. In contrast, insulin caused up to a 4‐fold decrease in the secretion rate of apolipoprotein B during the early growth periods. This inhibitory effect appeared to be partially abolished after 6 days. Our data suggest that some important questions on regulation of apolipoprotein B expression can be addressed by the long‐term culture of HepG2 cells in defined medium.
Identification of a receptor protein for neuropeptide Y in rabbit kidney G‐protein association and inhibition of adenylate cyclase
Authors: Gimpl Gerald , Wahl Jochen , Lang Rudolf E.
Published: July 2002

We previously identified the receptor for neuropeptide Y (NPY) on rabbit kidney membranes as a 100 kDa protein [(1990) J. Biol. Chem. 265, 18142–18143]. It is demonstrated in this study that the stable guanine nucleotide analogue, guanosine 5′‐O‐(3‐thiotriphosphate)(GTPγS), is capable of decreasing the labeling of the 100 kDa protein in a concentration‐dependent manner, with an IC20 value of 50 μM. This suggests that the 100 kDa protein represents a G‐protein coupled receptor type. In isolated proximal tubulex from rabbit kidney NPY decreases the parathyroid hormone stimulated cAMP production in a dose‐dependent fashion. This indicates that the NPY receptor in rabbit kidney is negatively coupled to adenylate cyclase by a G1‐like protein.
Actin depolymerization in the cyclic AMP‐stimulated toad bladder epithelial cell, determined by the DNAse method
Authors: Hays Richard M. , Lindberg Uno
Published: July 2002

Previous studies with the rhodamine phalloidin binding assay have shown that antidiuretic hormone and 8‐Br‐cAMP rapidly depolymerize F‐actin in toad bladder epithelial cells. We have extended these studies with DNAse inhibition assay and have found that in isolated epithelial cell suspensions, G‐actin increases from 37 to 56% of total actin following 8‐br‐cAMP stimulation. The G‐actin concentration in the epithelial cell greatly exceeds its critical concentration, indicating the requirement for a G‐actin sequestering protein or proteins in this system.
Cloning of a glycine receptor subtype expressed in rat brain and spinal cord during a specific period of neuronal development
Authors: Akagi Hiroyuki , Hirai Keiko , Hishinuma Fumio
Published: July 2002

Complementary (c) DNAs encoding a glycine receptor (GlyR) isomer were cloned from libraries constructed in λZAPII with poly (A)+ RNA of neonatal rat spinal cord. Northern blot analysis revealed that RNA hybridized to the cloned cDNA is detectable only for a period of late embryonic/ early postnatal stage of the spinal cord. Moreover, other central nervous tissues, such as hippocampus and cerebral cortex, in the infant rats are also rich in this message. The ‘neonatal (N) GlyR’ has 71% homology to that of another GlyR isoform in which adult rad cord is rich (AGlyR) Injection of a single RNA transcribed from the NGlyr‐cDNA into Xenopus oocyte induced functional formation of glycine‐gated Cl− channels, however, its pharmacological property differed from that of AGlyR.
Stoichiometry of metal‐tetracycline/H+ antiport mediated by transposon Tn10‐encoded tetracycline resistance protein in Escherichia coli
Authors: Yamaguchi Akihito , Iwasaki-Ohba Yoshiko , Ono Nobukazu , Kaneko-Ohdera Midori , Sawai Tetsuo
Published: July 2002

The tetracycline resistance protein (TetA) endoded by transposon Tn10 mediates the efflux of divalent cation‐tetracycline chelating complexes [Yamaguchi, A., Udagawa, T. and Sawai, T. (1990) J. Biol. Chem. 265, 4809–4813]. It was confirmed that protons were antiported with the complexes through an electrically‐neutral process because the antiport consumed ΔpH but not Δψ. The quantitative relationship between ΔpH and ΔpTC determined by a flow‐dialysis method clearly indicated a 1:1 stoichiometry of the monocationic metal‐tetracycline/H+ exchange.
Cyanobacteria contain a mitochrondrial complex I‐homologous NADH‐dehydrogenase
Authors: Berger Susanne , Ellersiek Ulrike , Steinmüller Klaus
Published: July 2002

Thylakoid and cytoplasmic membranes of the cyanobacterium Synechocystis sp. PCC 6803 were purified by sucrose gradient centrifugation. Both membranes oxidize NADH in a rotenone‐sensitive reaction. Antibodies prepared against psbG/ndhK and ndhJ fusion proteins detect the corresponding polypeptides in both membrane preparations. This demonstrates that a NADH‐dehydrogenase, homologous to the mitochondrial NADH‐ubiquinone‐oxidoreductase (complex I of the respiratory chain) is present in cyanobacteria. The NADH‐dehydrogenase can be solubilized with the detergent β‐D‐dodecylmaltoside. Sedimentation analysis of the solubilized enzyme on a sucrose gradient indicates that it is a multisubunit protein complex.
Occurrence of bifurcated three‐center hydrogen bonds in protpins
Authors: Preißner Robert , Egner Ursula , Saenger Wolfram
Published: July 2002

Analysis of 13 high‐resolution protein X‐ray crystal structures shows that 1204 (24%) of all the 4974 hydrogen bonds are of the bifurcated three‐center type with the donor X‐H opposing two acceptors A1, A2. They occur systematically in α‐helices where 90% of the hydrogen bonds are of this type; the major component is (n+4)N‐H…O=C(n) as expected for a 3.613 α‐helix, and the minor component is (n+4)N‐H…O=C(n+1), as observed in 310 helices; distortions at the C‐temini of α‐helices are stabilized by three‐center bonds, In β‐sheets 40% of the hydrogen bonds are three‐centered. The frequent occurrence of three‐center hydrogen bonds suggests that they should not be neglected in protein structural studies.
Association of a 24‐kDa GTP‐binding protein, Gn24, with human platelet α‐granule membranes
Authors: van der Meulen John , Bhullar Rajinder P. , Chancellor-Maddison Kerry-Ann
Published: July 2002

Human platelets were disrupted using nitrogen cavitation and fractionated on sucrose density gradients to permit isolation of α‐granules, the major secretory granule of platelets. Membrane proteins prepared from intact α‐granules by alkali extraction were separated by SDS‐polyacrylamide gel electrophoresis, transferred to nitrocellulose and the blot probed for the presence of GTP‐binding proteins using [α‐32P]GTP. Two low molecular mass GTP‐binding proteins with molecular mass of 27 and 24 kDa, respectively, were identified on the α‐granule membrane. In contrast to the 27‐kDa protein which was present in significant amounts in the plasma membrane‐enriched fraction, the 24‐kDa protein showed a preferential association with the α‐granule membrane. On immunoblotting with specific antiserum, the 24‐kDa GTP‐binding protein was found to be distinct from rab3A. To the best of our knowledge, the present report represents the first identification of low molecular mass GTP‐binding proteins associated with a platelet secretory granule.
‘Neuron‐specific’ protein gene product 9.5 (PGP 9.5) is also expressed in glioma cell lines and its expression depends on cellular growth state
Authors: Giambanco Ileana , Bianchi Roberta , Ceccarelli Paolo , Pula Grazia , Sorci Guglielmo , Antonioli Sabrina , Bocchini Virginia , Donato Rosario
Published: July 2002

Protein gene product 9.5 (PGP 9.5), which in the normal nervous system is restricted to certain neurons, has been detected in two glioma cell lines, rat C6 and human GL15, by immunoblotting and immunocytochemistry. Its expression in these cells depends on the cellular growth state, being maximal between the first and second post‐plating day. Only a faint PGP 9.5 immunoreactivity can be observed in glioma cells after the eleventh post‐plating day, i.e. about one week after confluency has been reached. The present results suggest that PGP 9.5 in cultured glial cells is maximally expressed during the growth phase and that the protein could play a role during brain development in glial cells, in reactive gliosis, or in tumorigenesis of the glial lineage.
Phosphorylation and inactivation of HMG‐CoA reductase at the AMP‐activated protein kinase site in response to fructose treatment of isolated rat hepatocytes
Authors: Gillespie John G. , Hardie D.Grahame
Published: July 2002

We have previously shown that incubation of isolated hepatocytes with fructose leads to elevation of AMP and activation of the AMP‐activated protein kinase. We now show that this treatment causes marked inactivation of HMG‐CoA reductase. Using immunoprecipitation from the microsomal fraction of 32P‐labelled cells, we also show that this treatment leads to a 2.6‐fold increase in the phosphorylation of the 100 kDa subunit of HMG‐CoA reductase. Successive digestion of this 32P‐labelled subunit with cyanogen bromide and endoproteinase Lys‐C confirmed that Ser‐871 the site phosphorylated in cell‐free assays by the AMP‐activated protein kinase, was the only site phosphorylated under these conditions.
Interaction of nitrogen dioxide with human plasma Antioxidant depletion and oxidative damage
Authors: Halliwell Barry , Hu Miao-Lin , Louie Samuel , Duvall Timothy R. , Tarkington Brian K. , Motchnik Paul , Cross Carroll E.
Published: July 2002

Nitrogen dioxide (NO*2) is often present in inhaled air and may be generated in vivo from nitric oxide. Exposure of human blood plasma to NO*2 caused rapid losses of ascorbic acid, uric acid and protein thiol groups, as well as lipid peroxidation and depletions of α‐tocopherol, bilirubin and ubiquinol‐10. No increase in protein carbonyls was detected. Supplementation of plasma with ascorbate decreased the rates of lipid peroxidation. α‐tocopherol depletion and loss of uric acid. Uric acid supplementation decreased rates of lipid peroxidation but not the loss of α‐tecopherol. We conclude that ascorbic acid, protein ‐SH groups, uric acid and α‐tocopherol may be important agents protecting against NO*2 in vivo. If these antioxidants are depleted, peroxidation of lipids occurs and might contribute to the toxicity of NO*2.
Implication of brain cdc2 and MAP2 kinases in the phosphorylation of tau protein in Alzheimer's disease
Authors: Ledesma M.D. , Correas I. , Avila J. , Diaz-Nido J.
Published: July 2002

Brain tau protein is phosphorylated in vitro by cdc2 and MAP2 kinases, obtained through immunoaffinity purification from rat brain extracts. The phosphorylation sites are located on the tau molecule both upstream and downstream of the tubulin‐binding motifs. A synthetic peptide comprising residues 194–213 of the tau sequence, which contains the epitope recognized by the monoclonal antibody tau‐1, is also efficiently phosphorylated in vitro by cdc2 and MAP2 kinases. Phosphorylation of this peptide markedly reduces its interaction with the antibody tau‐1, as it has been described for tau protein in Alzheimer's disease. Both cdc2 and MAP2 kinases are present in brain extracts obtained from Alzheimer's disease patients. Interestingly, the level of cdc2 kinase may be increased in patient brains as compared with non‐demented controls. These results suggest a role for cdc2 and MAP2 kinases in phosphorylating tau protein at the tau‐1 epitope in Alzheimer's disease.
Flow‐induced prostacyclin production is mediated by a pertussis toxin‐sensitive G protein
Authors: Berthiaume Francois , Frangos John A.
Published: July 2002

Fluid flow and several other agonists induce prostacyclin (PGI2) production in endothelial cells, G proteins mediate the response of a large number of hormones such as histamine, but the transduction pathway of the flow signal is unclear. We found that GDPβS and pertussis toxin inhibited flow‐induced prostacyclin production in human umbilical vein endothelial cells. In addition, flow potentiated the histamine‐induced production of PGI2. This suggests that flow stimulates prostacyclin production via a pertussis toxin‐sensitive G protein and modulates the stimulus‐response coupling of other agonists.
Regulatory regions in the yeast FBP1 and PCK1 genes
Authors: Mercado Juan J. , Gancedo Juana M.
Published: July 2002

By deletion analysis of the fusion genes FBP1‐lacZ and PCK1‐lacZ we have identified a number of strong regulatory regions in the genes FBP1 and PCK1 which encode fructose‐1,6‐bisphosphatase and phosphoenolpyruvate carboxykinase. Lack of expression of β‐galactosidase in fusions lacking sequences from the coding regions suggests the existence of downstream activating elements. Both promoters have several UAS and URS regions as well as sites implicated in catabolite repression. We have found in both genes consensus sequences for the binding of the same regulatory proteins, such as yAP1, MIG1 or the complex HAP2/HAP3/HAP4. Neither deletion nor overexpression of the MIG1 gene affected the regulated expression of the FBP1 or PCK1 genes.
The catalytic activities of monomeric enzymes show complex pressure dependence
Authors: Groβ Michael , Auerbach Günter , Jaenicke Rainer
Published: July 2002

High hydrostatic pressures in the biologically relevant range (⩽ 1,200 bar) are known to cause dissociation of oligomeric enzymes in vitro, whereas protein denaturation requires pressures far beyond this range. Pressure‐induced inactivation phenomena attributable to neither of these effects are shown to occur in monomeric enzymes. Three different types of pressure dependence can be distinguished: (1) a linear dependence of catalytic rate constants on pressure, as predicted by the activated complex theory, observed for lysozyme and thermolysin; (2) a biphasic profile consisting of two linear contributions, found for trypsin; (3) maximum curves, as observed for both directions of the octopine dehydrogenase reaction. The third case may be ascribed to a pressure‐induced decrease in the partial specific volume of the protein, resulting in reduced flexibility of the active site. This mechanism may also apply to the pressure‐induced inactivation of assembly systems stabilized against dissociation in the cell.
Molecular mechanism of inhibition of mammalian protein synthesis by some four‐chain agglutinins
Authors: Citores Lucía , Ferreras J.Miguel , Iglesias Rosario , Carbajales Mercedes L. , Arias F.Javier , Jiménez Pilar , Rojo M.Angeles , Girbés Tomás
Published: July 2002

The four chain agglutinins from Abrus precatorius, Viscum album and Ricinus communis promote depurination of the 28 S rRNA from rabbit reticulocyte ribosomes characteristic of the common ribosome‐inactivating proteins (RIPs). These agglutinins inhibited mammalian protein synthesis at nanomolar concentrations but they do not affect plant protein synthesis under the same conditions. Therefore, they should also be considered as true RIPs but of a new class, the four‐chain RIPs. An extended classification of RIPs is presented based on the former one from Stirpe et al. [Bio/technology 10 (1992) 405‐412].
Generation of Superoxide by purified and relipidated cytochrome b559 in the absence of cytosolic activators
Authors: Koshkin Vasilij , Pick Edgar
Published: July 2002

Purified cytochrome b559
from guinea pig macrophages was relipidated with several phospholipid mixtures. Relipidated cytochrome b559
was found capable of NADPH‐dependent Superoxide (O2
−) production in the absence of the cytosolic components of the NADPH oxidase complex. The rate of O2
− generation by cytochrome b559
varied with the type of phospholipid utilized for relipidation, was absolutely dependent on exogenous FAD, and was enhanced by a critical concentration of anionic amphiphile. It is demonstrated that exogenous FAD acts by binding to cytochrome b559
. These results provide firm experimental evidence for the proposal that cytochrome b559
comprises the complete electron transporting apparatus of the O2
− forming NADPH oxidase and that the cytosolic components function merely as activators.
The point mutation of mitochondrial DNA characteristic for MERRF disease is found also in healthy people of different ages
Authors: Münscher C. , Rieger T. , Müller-Höcker J. , Kadenbach B.
Published: July 2002

The A‐to‐G transition mutation in the tRNALys gene of mitochondrial DNA (mtDNA), characteristic for the maternally inherited MERRF syndrome (myoclonic epilepsy with ragged red fibers), has been identified by point mutation‐specific polymerase chain reaction in extraocular muscle from 11 of 16 healthy people of different ages. No mutation was found in navel‐string samples from 5 newborns, in HeLa cells, and in 2 individuals younger than 20 years. On the other hand, the mutation is present in all 5 tested 74–89‐year‐old individuals and in 6 of 9 20–70‐year‐old individuals. The amount of mutated from total mtDNA was estimated by ‘mispairing PCR’ in extraocular muscle of 2 individuals of 74 and 89 years to 2.0 and 2.4%, respectively. In most tissue samples the MERRF mutation occurs together with the ‘common deletion’ of mtDNA, which was previously shown to accumulate in healthy individuals with increasing age. It is proposed that during aging, deletions and point mutations of mtDNA accumulate, which could impair mitochondrial energetics.
Identification of protein phosphatase 2A as the primary target for microcystin‐LR in rat liver homogenates
Authors: Toivola Diana M. , Eriksson John E. , Brautigan David L.
Published: July 2002

The liver‐specific toxin microcystin‐LR (MC‐LR) is a potent inhibitor of type 1 (PP1) and type 2A (PP2A) protein phosphatases. A tritiated form of the toxin, [3H]dihydromicrocystin‐LR ([3H]DMC‐LR), was used to identify target proteins in cellular fractions prepared from rat liver homogenates. About 80% of the [3H]DMC‐LR bound to proteins was in the cytosolic fraction, which contained essentially all of the PP2A. In contrast, much of the PP1 was found in particulate fractions, each with only a few percent of the total protein‐bound [3]HDMC‐LR. Protein‐bound [3H]DMC‐LR in the cytosol co‐eluted with PP2A, but not with PP‐1 from a DEAE‐Sepharose column. Native forms of liver cytoplasmic PP2A and PP1 separated by aminohexyl‐Sepharose adsorption showed similar sensitivity to inhibition by MC‐LR, and bound [3H]DMC‐LR proportional to the amount of phosphatase activity. The results indicate that [3H]DMC‐LR can bind both PP2A and PP1 in the liver which must be important for microcystin‐induced toxicity, but is recovered mainly bound to PP2A in the cytosol.