bluuebunny/crossref_metadata_2025 huggingface dataset:
A Simple Risk Scoring System for Predicting Recurrence in Women with Locally Advanced Breast Cancer (LABC) Treated with Neoadjuvant Chemotherapy (NACT)
Authors: Agrawal Sushma , Mishra Prabhakar , Mishra Anjali , Agarwal Gaurav
Published: June 2021

Introduction: Breast cancer commonly presents in locally advanced stage (LABC) in developing countries, for which NACT followed by surgery and radiotherapy is the standard of care. There is a need for a simple tool to risk categorise patients in the clinic, so that treatment intensification can be offered to women with high risk of recurrence.

Materials and Methods: Data of prospectively maintained database of LABC (between January 2007 - December 2012), who received NACT followed by surgery, radiotherapy and endocrine therapy was retrospectively analysed for clinico-pathological factors associated with disease recurrences. A recurrence risk scoring model was developed on the basis of regression coefficient of identified independent risk factors.

Results: In the data set of 206 patients, the median follow-up was 48 months (range: 6-156 months) and mean and median disease-free survival (DFS) were 87.41 and 85 months. The 1, 5, 10 years DFS was 95%, 54% and 41%. The independent risk factors (on modified p value <0.40) for recurrence were Tumour stage, Nodes stage, grade, age groups, pathologic complete response, intrinsic subtype, and type of surgery. Risk score prepared by regression coefficient (β), was in the range of 1-8 with median score of 5. ROC curve showed that area under ROC Curve of the score was 71.8% (95% CI: 64.8%-78.8%, p<0.001). To detect recurrences, a risk score ≥3 had at least 93.1% sensitivity and 31.9% specificity whereas score ≥4 had at least 73.5% sensitivity and 59.6% specificity. Based on cluster analysis, score 1-4 was identified as low risk whereas 5-6 as moderate risk group and ≥7 identified as high-risk group and their mean/median disease free survival time were 107.86/ NR, 66.99/30 months and 58.34/20 months respectively.

Conclusions: The significant difference in DFS among three risk groups, indicates goodness of the fit of our risk score model. The risk scoring model developed by us is simple, easy to use in clinic and can be used for selecting high risk patients who benefit from treatment intensification.
Using Glufosinate in Sequential Application Systems with Dicamba on Palmer Amaranth
Authors: Ogden Grace Flusche , Dotray Peter A.
Published: June 2021

Aims: Determine the influence of sequential spray order and role of glufosinate when used in a system with dicamba to control Palmer amaranth at three different growth stages.

Study design:  Randomized complete block design with four replications

Place and duration of Study: A fallow, non-crop field at the Texas A&M AgriLife Research and Extension Center, Lubbock, Texas, during the 2018-2019 growing seasons

Methodology: Herbicides were applied to < 10 cm, 10 to 20 cm, and > 30 cm Palmer amaranth with a handheld 1.93m CO2-pressurized backpack calibrated to deliver 140 L ha-1 at 207 kPa. Palmer amaranth control was visually estimated on a scale of 0 (no control) to 100% (complete plant death) relative to the nontreated control. Palmer amaranth biomass and density were taken when all plots reached 50% or less control in 2019. Palmer amaranth control, biomass, and density were subjected to analysis of variance and means were separated using Fisher’s Protected LSD at an alpha of 0.05.

Results: Palmer amaranth control decreased as Palmer amaranth size at initial application increased. A difference in efficacy based on herbicide order was observed for < 10 cm Palmer amaranth. Glufosinate followed by dicamba was less effective (76-83%) than dicamba followed by glufosinate (93-96%) at 2 of 4 rating dates following sequential applications in both years. Dicamba + acetochlor followed by glufosinate provided greater Palmer amaranth control than dicamba followed by dicamba at one or more rating dates across all weed sizes.

Conclusion: Glufosinate served as a complimentary partner in the dicamba-based system, and additional modes of action will be more effective to slow the development of resistance to group 4 herbicides when compared to repeated use of a group 4 herbicide used alone.
Study of Hydrolysis Process from Pineapple Leaf Fibers using Sulfuric Acid, Nitric Acid, and Bentonite Catalysts
Authors: Wiyantoko Bayu , Rusitasari Rika , Putri Rahma Novia
Published: June 2021

The hydrolysis process of pineapple leaf fibers has been carried out using sulfuric acid, nitric acid, bentonite catalyst, and activated bentonite catalyst. The sugar content of the hydrolysis product was identified using the phenol-sulfuric acid method by UV-Visible spectrophotometry. The disposal of pineapple leaf is a big problem even though it has high cellulose content (70–80%) and very promising to produce sugar by hydrolysis process. The purpose of this experiment was to determine the effectiveness of homogeneous and heterogeneous catalysts related to sugar levels in pineapple leaf fiber. The variables in this study were the sampling time during the hydrolysis process at a temperature of 100 °C and the addition of homogeneous and heterogeneous catalysts. The homogeneous catalysts were sulfuric acid and the nitric acid meanwhile heterogeneous catalyst was thermally activated bentonite and acid-activated bentonite. The results obtained highest sugar content reached at 150 minutes using chemical activated bentonite catalysts at 6.459 g/L and the addition of catalysts affected sugar yields, speed up the reaction, bentonite as a good catalyst, and gave good prospect for ethanol production in further process. Copyright © 2021 by Authors, Published by BCREC Group. This is an open access article under the CC BY-SA License (https://creativecommons.org/licenses/by-sa/4.0).
Challenges & Opportunities on Catalytic Conversion of Glycerol to Value Added Chemicals
Authors: Zakaria Zaki Yamani , Jusoh Mazura , Kader Shams Shazid , Idris Siti Shawalliah
Published: June 2021

With the rapid expansion of biodiesel industry, its main by-product, crude glycerol, is anticipated to reach a global production of 6 million tons in 2025. It is actually a worrying phenomenon as glycerol could potentially emerge as an excessive product with little value. Glycerol, an alcohol and oxygenated chemical from biodiesel production, has essentially enormous potential to be converted into higher value-added chemicals. Using glycerol as a starting material for value-added chemical production will create a new demand on the glycerol market such as lactic acid, propylene glycol, alkyl lactatehydrogen, olefins and others. This paper briefly reviews the recent development on value-added chemicals derived from glycerol through catalytic conversion of refined and crude glycerol that have been proven to be promising in research stage with commercialization potential, or have been put in a corporate marketable production. Despite of the huge potential of products that can be transformed from glycerol, there are still numerous challenges to be addressed and discussed that include catalyst design and robustness; focus on crude or refined glycerol; reactor technology, reaction mechanism and thermodynamic analysis; and overall process commercial viability. The discussion will hopefully provide new insights on justified direction to focus on for glycerol transformation technology. Copyright © 2021 by Authors, Published by BCREC Group. This is an open access article under the CC BY-SA License (https://creativecommons.org/licenses/by-sa/4.0).
Synthesis of Ash Derived Co/Zeolite Catalyst for Hydrogen Rich Syngas Production via Partial Oxidation of Methane
Authors: Zaffar Amer , Khan Bilal Alam , Khoja Asif Hussain , Khan Uneeb Masood , Sarmad Qassam , Mehran Muhammad Taqi , Naqvi Salman Raza , Ali Majid
Published: June 2021

The objective of this study was to analyze the catalytic performance of series of cobalt-modified Zeolite-4A supported catalysts for the syngas (CO and H2) production at 800 °C via the partial oxidation of methane (POM). The Co/Zeolite-4A catalyst was synthesized using a two-step hydrothermal method from coal fly ash. The synthesized catalysts were characterized by X-ray Diffraction (XRD), Scanning Electron Microscope (SEM), Energy Dispersive X-ray (EDX), and Thermogravimetric Analysis (TGA). The catalyst shows a crystalline structure with stability up to 900 °C. The catalytic performance analysis shows the CH4 conversion increases from 29 to 68% for 0 and 10 wt% Co over Zeolite-4A, respectively. The H2 selectivity was improved from 28–56% while CO selectivity increased from 24–52 % making H2/CO ratio > 1. The stability analysis shows the 10% Co/Zeolite-4A withstand for 24 h a time on stream (TOS). Finally, the spent catalyst analysis was carried out to check the carbon formation along with its structural analysis. The minimal carbon formation is analyzed in 24 h TOS for POM reaction. Copyright © 2021 by Authors, Published by BCREC Group. This is an open access article under the CC BY-SA License (https://creativecommons.org/licenses/by-sa/4.0).
Encapsulation of hemoglobin in phospholipid vesicles
Authors: Gaber Bruce P. , Yager Paul , Sheridan James P. , Chang Eddie L.
Published: July 2002

Hemoglobin has been encapsulated in phospholipid vesicles by extrusion of hemoglobin/lipid mixtures through polycarbonate membranes. This technique avoids the use of organic solvents, sonication, and detergents which have proven deleterious to hemoglobin. The vesicles are homogeneous, with a mean size of 2400 Å as determined by photon correlation spectroscopy. The encapsulated hemoglobin binds oxygen reversibly and the vesicles are impermeable to ionic compounds. Hemoglobin encapsulated in egg phosphatidylcholine vesicles converts to methemoglobin within 2 days at 4°C. By contrast, when a mixture of dimyristoyl phosphatidylcholine, cholesterol and dicetyl phosphate is used there is no acceleration in methemoglobin formation, and the preparation is stable for at least 14 days at 4°C.
Reticulocyte lipoxygenase exhibits both n‐6 and n‐9 activities
Authors: Kühn H. , Wiesner R. , Schewe T. , Rapoport S.M.
Published: July 2002

Purified reticulocyte lipoxygenase converts arachidonic acid to both 15‐ and 12‐hydroxyperoxyeicosatetraenoic acids. The proportion of the two reaction products does not change during the purification procedure as shown by HPLC analysis. By means of isoelectric focusing it was not possible to separate the n‐6 and n‐9 activities. Reticulocyte lipoxygenase was completely inactivated by both 5,8,11‐eicosatriynoic and 5,8,11,14‐eicosatetraynoic acids in contrast to soybean lipoxygenase‐1 which inactivated only by 5,8,11,14‐eicosatetraynoic acid. These results indicate that reticulocyte lipoxygenase exhibits both n‐6 and n‐9 activities. A contamination of the enzyme preparation with other lipoxygenases, e.g., the n‐9 lipoxygenase from thrombocytes appears to be excluded.
The variant surface glycoproteins of Trypanosoma equiperdum
Authors: Baltz T. , Duvillier G. , Giroud Ch. , Richet C. , Baltz D. , Degand P.
Published: July 2002

The cross‐reacting antigenic determinant in the variant surface glycoproteins (VSGs) of Trypanosoma equiperdum was studied by testing the ability of VSG glycopeptides to bind heterologous anti‐VSG sera. VSG glycopeptide purification revealed the presence of 3 oligosaccharide sidechains on the mature VSG. These consist of two sidechains containing only mannose and glucosamine and a third containing galactose and mannose (in a 5 : 1 ratio) as well as phosphorous and ethanolamine. This phosphorylated fragment completely blocked the binding of VSG to heterologous anti‐VSG and therefore contained the cross‐reacting determinants.
Attachment of origins of replication to the nuclear matrix and the chromosomal scaffold
Authors: van der Velden Huub M.W. , van Willigen Gijsbert , Wetzels Ria H.W. , Wanka Friedrich
Published: July 2002

We have investigated the attachment of DNA to the nuclear matrix and chromosomal scaffold in synchronized bovine liver cells. Label incorporated at the onset of the S phase remained preferentially associated with the matrix during the subsequent G1 phase and with a residual protein structure from dehistonized chromosomes during mitosis. On the other hand label incorporated during mid or late S phase was about equally distributed over the DNA molecule after a chase into the G1 phase. These results suggest that DNA is attached to the nuclear matrix and chromosome scaffolds by the origins of replication.
Activation of rat heart phosphofructokinase‐2 by insulin in vivo
Authors: Rider Mark H. , Hue Louis
Published: July 2002

Fifteen minutes after the intravenous injection of overnight starved, anaesthetized rats with insulin, the concentration of fructose 2,6‐bisphosphate was increased more than 2‐fold in the hearts of these animals. Insulin injection also caused a 2–3‐fold increase in the V
maxof phosphofructokinase‐2 with no detectable change in K
m values. The effect persisted after precipitation of the enzyme with polyethylene glycol or after gel filtration through Sephadex G‐25.
Phorbol ester inhibits myoblast fusion and activates β‐adrenergic receptor coupled adenylate cyclase
Authors: Sulakhe Prakash V. , Johnson Dennis D. , Phan Ngoc T. , Wilcox Robert
Published: July 2002

Primary cultures of myoblasts, derived from embryonic chick pectoral muscle, were treated with phorbol ester (TPA) for 8–96 h. TPA treatment blocked the fusion of myoblasts along with the expression of the MM form of creatine kinase. Interestingly, TPA treatment markedly increased the activity of β‐adrenergic receptor coupled adenylate cyclase (AC) activity. The study suggests that TPA treatment augments the functional interaction between a coupling Ns protein and catalytic unit of AC. The likely significance of these results is briefly presented.
Pentose cycle pathway in normal and tumoral islet cells
Authors: Giroix Marie-Hélène , Sener Abdullah , Malaisse Willy J.
Published: July 2002

Relative to protein content, the activity of glucose‐6‐phosphate dehydrogenase and 6‐phosphogluconate dehydrogenase and the rate of glucose metabolism by the pentose cycle pathway in tumoral insulin‐producing cells were similar to or higher than those found in normal rat islets. Hence, the decreased secretory response of tumoral cells to glucose is apparently not attributable to any major anomaly in glucose handling by the hexose monophosphate pathway.
Interaction of clathrin with liposomes: pH‐dependent fusion of phospholipid membranes induced by clathrin
Authors: Hong Keelung , Yoshimura Tetsuro , Papahadjopoulos Demetrios
Published: July 2002

Clathrin, dissociated from coated vesicles of bovine brain and purified by gel chromatography, was found to interact with the lipid bilayer as shown by the spontaneous release of encapsulated fluorescent dye in liposome. Clathrin‐induced dye release was enhanced at acidic pH in phosphatidylserine‐containing vesicles. A strong correlation between dye release and fusion of liposomes was observed. In general, when there was a fast release of encapsulated dye induced by clathrin, a pH‐dependent, clathrin‐induced fusion was observed. Clathrin did not induce either dye release or fusion of egg phosphatidylcholine liposomes. The self‐association of clathrin at low pH diminished the fusogenic activity. Fusion induced by clathrin at low pH c̀ould be stopped at pH above 5.0 and resumed by lowering the pH below 5.0. This suggests that the interaction of clathrin with phospholipid membranes can be regulated by pH.
Nature of carbon monoxide binding of iron(II)‐bleomycin
Authors: Sugiura Yukio , Kuwahara June , Suzuki Tadashi
Published: July 2002

Iron‐bleomycin
CO affinity
Photolysis
Steric effect
Occurrence of lipid and phorbol ester activated protein kinase in wheat cells
Authors: Oláh Zoltán , Kiss Zoltán
Published: July 2002

Chromatography of the 30000 × g supernatant of wheat cell homogenates on DEAE‐cellulose yielded a fraction whose protein kinase activity was specifically stimulated by phosphatidylserine plus phorbol ester or phosphatidylserine plus diolein. Such a characteristic activation pattern established this protein kinase as being a kinase C, an enzyme which plays an important role in the regulation of cell growth and differentiation in animal cells.
Interactions of tuftsin with bovine serum albumin
Authors: Bhakuni Vinod , Gupta C.M.
Published: July 2002

Interactions of tuftsin (Thr‐Lys‐Pro‐Arg) with bovine serum albumin (BSA) were analysed by fluorescence spectroscopy and circular dichroism. The data show that tuftsin interacts weakly with BSA, but this interaction is considerably enhanced by introducing an apolar substituent at the C‐terminus of the tetrapeptide. It is suggested that strong binding of tuftsin to albumin in blood may enhance its macrophage‐stimulating activity in vivo.
The polar head group of a novel insulin‐sensitive glycophospholipid mimics insulin action on phospholipid methyltransferase
Authors: Kelly Kathleen L. , Mato Jose M. , Jarett Leonard
Published: July 2002

A phospholipid has been purified from rat liver membranes which copurified with an insulin‐sensitive glyco‐phospholipid isolated from H35 hepatoma cells. The polar head group of this phospholipid was generated by treatment with a phosphatidylinositol‐specific phospholipase C from Staphylococcus aureus and purified through a C18 extraction column. Like insulin, the addition of this polar head group to isolated rat adipocytes inhibited the stimulatory effect of isoproterenol on phospholipid methyltransferase. The polar head group was also active on a subcellular fraction. The addition of the polar head group to microsomes isolated from isoproterenol‐treated adipocytes produced a time‐dependent inactivation of phospholipid methyl‐transferase, approaching basal activity. It is proposed that the effects of insulin on phospholipid methyl‐transferase may be mediated by this polar head group.
The stimulation of phosphorylation of intracellular proteins in GH3 rat pituitary tumour cells by phorbol esters of distinct biological activity
Authors: Brooks S.F. , Evans F.J. , Aitken A.
Published: July 2002

Using a pituitary tumour cell line (GH3), we have studied the phosphorylation of intracellular proteins induced by phorbol esters of diverse biological activity. All the active phorbol esters, including the weakly tumour‐promoting but non‐platelet aggregatory compound DOPPA, stimulated the phosphorylation of a cytosolic 80 kDa protein. A protein of this molecular mass has been suggested to be a marker of PKC activity. In contrast, only TPA and the non‐tumour promoting but highly active phorbol ester SAP A stimulated the phosphorylation of a 130 kDa membrane protein. The results suggest that these phorbol esters activate PKC, but induce the differential phosphorylation of a variety of intracellular proteins.
Binding of low‐density lipoprotein to monolayer cultures of rat hepatocytes is increased by insulin and decreased by dexamethasone
Authors: Salter Andrew M. , Fisher Sylvia C. , Brindley David N.
Published: July 2002

Rat hepatocytes were maintained in monolayer culture for 20 h in the presence of 10% (v/v) new‐born calf serum and then for a further 1–24 h in serum‐free medium containing 2 g bovine serum albumin/l. The specific binding of human 125‐I‐LDL to two distinct sites was then measured at 4°C. Binding to site 1 was displaced by dextran sulphate while that to site 2 was not. The presence of 1–100 nM insulin for 24 h in the second incubation significantly increased binding to site 1. Significant increases were also seen when cells were incubated with 10 nM insulin for 1 h. No significant effects of insulin on binding to site 2 were observed. In contrast, 10 nM—1 μM dexamethasone decreased binding to both sites. The effects of these hormones were mutually antagonistic.
Increased membrane heterogeneity in stimulated human granulocytes
Authors: Valentino Matteo , Governa Mario , Gratton Enrico , Fiorini Rosamaria , Curatola Giovanna , Bertoli Enrico
Published: July 2002

TMA‐DPH fluorescence decay in human PMN before and after stimulation with FMLP was studied using frequency domain fluorometry. Membrane heterogeneity was assessed by the width of the continuous distributions of lifetime values of Lorentzian shape used to describe the fluorescence decay. In non‐stimulated granulocytes TMA‐DPH fluorescence decay is characterized by two distributions of lifetime values centered at 6.5 and 1.0 ns and full width at half maximum of 0.3 and 1.2 ns, respectively. Within 15 min after stimulation, the center values of the two distribution components were 5.1 and 0.8 ns and the distribution width was 0.8 and 0.6 ns, respectively. These results indicate changes of membrane domain organization which can be ascribed to compositional changes redistribution of membrane components.
Heat‐stable microtubule protein MAP‐1 binds to microtubules and induces microtubule assembly
Authors: Vera Juan C. , Rivas Coralia I. , Maccioni Ricardo B.
Published: July 2002

The microtubule‐associated proteins, MAP‐1, MAP‐2 and tau, have been purified from brain tissue via a new approach using a heating step directly on the homogenate, followed by selective adsorption on calmodulin‐Sepharose affinity columns and gel‐filtration chromatography. Our results indicate that these MAPs share common biochemical properties, including heat stability, calmodulin binding and promotion of tubulin assembly into microtubules.
Insulin‐induced decrease in 5′‐nucleotidase activity in skeletal muscle membranes
Authors: Klip Amira , Ramlal Toolsie , Douen Andre G. , Burdett Elena , Young Douglas , Cartee Gregory D. , Holloszy John O.
Published: July 2002

Insulin releases inositol phosphoglycans from myocytes in culture [(1986) Science 233, 967–972], which display insulinomimetic activity. Because 5′‐nucleotidase is anchored to the membrane through inositol‐containing phospholipid glycans, we investigated whether insulin could release the enzyme from the membrane. Membranes prepared from hindquarter muscles of rats perfused with insulin showed a 23% decrease in 5′‐nucleotidase activity. Isolated membranes from muscle exposed to insulin in vitro also showed a small but reproducible decrease (9%) in 5′‐nucleotidase activity relative to unexposed controls. Phospholipase C from Staphylococcus aureus released 60% of the membrane‐bound 5′‐nucleotidase. We propose that insulin may activate an endogenous phospholipase C that cleaves phospholipid‐glycan‐anchored proteins.
Single amino acid substitution between SHV‐1 β‐lactamase and cefotaxime‐hydrolyzing SHV‐2 enzyme
Authors: Barthélémy Michel , Péduzzi Jean , Ben Yaghlane Hasna , Labia Roger
Published: July 2002

SHV‐2 β‐lactamase was purified from an overproducing variant of a clinical isolate of Escherichia coli resistant to cefotaxime. Pure protein was digested by trypsin and Lys‐C endoproteinase. Proteolytic peptides, isolated by reverse‐phase HPLC, were submitted to manual Edman degradation and aligned by homology with the sequence of SHV‐1 β‐lactamase. A putative amino acid sequence was deduced. Structural comparison revealed that SHV‐2 differed from SHV‐1 by only one amino acid, Gly → Ser, at position 213 of the mature protein.
Structural requirements of choline derivatives for ‘conversion’ of pneumococcal amidase A new single‐step procedure for purification of this autolysin
Authors: Sanz Jesus Miguel , Lopez Rubens , Garcia Jose Luis
Published: July 2002

Tertiary amines appear to be the minimal structure needed to convert in vitro the inactive form (E‐form) of pneumococcal amidase to the catalytic active form (C‐form). Diethylethanolamine was one of the compounds that converted the E‐form, a finding that has been used successfully to develop an affinity chromatography system in DEAE‐cellulose for the rapid and efficient purification of lytic enzymes of pneumococcus and its bacteriophages.
Structure of the 5′‐external transcribed spacer of the human ribosomal RNA gene
Authors: Renalier Marie-Hélène , Mazan Sylvie , Joseph Nicole , Michot Bernard , Bachellerie Jean-Pierre
Published: July 2002

We report the complete nucleotide sequence of the 3627 bp long 5′‐external transcribed spacer (ETS) of a human ribosomal RNA gene. This sequence exhibits only very limited homologies with its mouse counterpart, the only other mammalian specimen analyzed so far. It has very peculiar compositional characteristics, with a highly biased base content (very rich in G+C, very poor in A) and also some very strong dinucleotide preferences. Interestingly, these specific features are shared by the mouse sequence, despite the extensive sequence divergence, and also apply to the other transcribed spacers of mammals indicating that a common and strong structural constraint is exerted on all these regions of the ribosomal gene. An outstanding secondary structure can be formed within the human ETS RNA, which could have a significant role in preribosome assembly.
Fusion to an endoglucanase allows alkaline phosphatase to bind to cellulose
Authors: Greenwood Jeffrey M. , Gilkes Neil R. , Kilburn Douglas G. , Miller Robert C. , Warren R.Antony J.
Published: July 2002

Endoglucanase CenA of Cellulomonas fimi comprises an N‐terminal cellulose‐binding domain and a C‐terminal catalytic domain joined together by a sequence of 23 proline and threonine residues (the Pro‐Thr box). The domains function independently when separated by proteolysis. TnphoA has been used to generate cenA′‐′phoA fusions. CenA′‐′PhoA fusion polypeptides which contain the entire cellulose‐binding domain of CenA bind to cellulose, allowing their purification from periplasmic extracts in a single, facile step. This result has implications for purification or immobilisation of chimeric proteins on a cheap cellulose matrix.
Presence of immunoreactive endothelin in human milk
Authors: Lam Hing-Chung , Takahashi Kazuhiro , Ghatei Mohammed A. , Bloom Stephen R.
Published: July 2002

Endothelin‐like immunoreactivity was detected in human milk at a concentration of 6.8 ± 1.6 (mean ± SEM; n = 16) using a highly sensitive radioimmunoassay. Gel filtration and fast protein liquid chromatography (FPLC) verified the identity of the endothelin. FPLC revealed 4 peaks, one eluting just after the void volume, and the other three in the positions of endothelin‐1, ‐2, and ‐3, respectively.
Copolymers of glutamic acid and tyrosine are potent inhibitors of oocyte casein kinase II
Authors: Tellez Rowena , Gatica Marta , Allende Catherine C. , Allende Jorge E.
Published: July 2002

Polypeptides rich in glutamic acid are strong inhibitors purified from isolated nuclei of Xenopus laevis oocytes of casein kinase II. The presence of tyrosine in these peptides greatly enhances their inhibitory capacity. Using casein as a substrate, copolyglu:tyr (4:1) has an I
50 value of 20 nM, 250 fold lower than that of polyglutamic acid which is 5 μM. A similar large difference is observed when a synthetic peptide is used as substrate. The inhibition of copolyglu:tyr is competitive with casein and can be completely reversed by high ionic strength. The relative inhibitory capacity of the polypeptides tested, in descending order, is copolyglu:tyr (4:1) > copolyglu:tyr (1:1) > polyglu > copolyglu:phe (4:1) > copolyglu:ala ( > copolyglu:leu (4:1). The high affinity for tyrosine‐containing acid peptides is shared by rat liver and yeast casein kinase II so that it seems to be a general property of these enzymes.
Dual regulation by G proteins of agonist‐dependent phosphorylation of muscarinic acetylcholine receptors
Authors: Haga Kazuko , Haga Tatsuya
Published: July 2002

Muscarinic acetylcholine receptors purified from porcine atrium were phosphorylated, depending on the presence of agonists, by a protein kinase partially purified from porcine brain, which had similar properties to the β‐adrenergic receptor kinase. GTP‐binding regulatory proteins (Go) had dual effects on the phosphorylation of muscarinic receptors, i.e. stimulation at lower concentrations and inhibition at higher concentrations. The stimulatory effect was reproduced with the βγ subunit of Go and the inhibitory effect with the combination of the α and βγ subunits.
RNA editing of ATPase subunit 9 transcripts in Oenothera mitochondria
Authors: Schuster Wolfgang , Brennicke Axel
Published: July 2002

The mRNA for subunit 9 of the ATPase (atp9) in the higher plant Oenothera is edited in four nucleotide positions. Three events alter genomic serine and proline codons to triplets specifying leucine. A UGA termination codon is introduced into the reading frame by modification of a CGA arginine codon. This modification shortens the polypeptide by four amino acids. Direct sequencing of PCR amplified cDNA from the total mitochondrial mRNA population gives no indication of partially edited transcripts suggesting a rapid and efficient modification of atp9 transcripts in Oenothera mitochondria.
Site‐directed mutagenesis of La protease
Authors: Amerik Alexander Yu. , Antonov Vladimir K. , Gorbalenya Alexander E. , Kotova Svetlana A. , Rotanova Tatyana V. , Shimbarevich Elena V.
Published: July 2002

Comparative sequence analysis of Escherichia coli ATP‐dependent La protease led to the suggestion that Ser679 is the catalytically active enzyme residue. Site‐directed mutagenesis Ser679 → Ala, investigation of the cells containing the mutant plasmid, and study of the partially purified mutant protein produced results in favour of this suggestion.
Calponin Developmental isoforms and a low molecular weight variant
Authors: Draeger A. , Gimona M. , Stuckert A. , Celis J.E. , Small J.V.
Published: July 2002

Two‐dimensional gel analysis of basic proteins in developing human smooth muscle identifies calponin as a prominent marker of the differentiated phenotype. Adult tissue (human and mouse) typically expresses up to four calponin isoforms, three of which appear sequentially during fetal development: adult myometrial cells express the same three isoforms in primary culture in vitro and these are down‐regulated, in reverse order, during the subsequent modulation of phenotype. Monospecific, polyclonal antibodies against calponin identify a lower molecular weight variant of calponin (L‐calponin) that is strongly and specifically expressed in adult smooth muscles of the human urogenital tract. L‐Calponin is down‐regulated in benign smooth muscle derived tumors (leiomyoma) and is not expressed in primary cultures of normal uterine tissue.
POMC gene expression in mouse and hamster melanoma cells
Authors: Slominski Andrzej
Published: July 2002

Expression of mRNA hybridizable to murine POMC cDNA, and immunoreactivity specific for POMC derived peptide, β‐endorphin, are reported for the first time in one murine and two hamster melanoma lines. The apparent sizes of POMC mRNA transcript were 3.5, 1.5 and 1 kb. It is suggested that POMC gene expression may be involved in the autoregulation of the melanoma phenotype at the cellular and tissue levels.
Probable occurrence of toxin‐susceptible G proteins in the nematode Caenorhabditis elegans
Authors: Arata Yoichiro , Tada Shusuke , Ui Michio
Published: July 2002

Pertussis toxins, islet‐activating protein (IAP), and cholera toxin ADP‐ribosylated 40 kDa and 45 kDa proteins in membrane preparations from Caenorhabditis elegans. Proteins with the same molecular weights were recognized in the same membranes by an antibody that had been raised against a peptic common to α‐subunits of mammalian αβγ‐heterotrimeric G proteins. The antibody produced immunoprecipitation with the 40 kDa protein 32P‐labeled by IAP, A 35 kDa protein immunochemically indistinguishable from β‐component of mammalian G proteins was also found in C. elegans membranes. The membranes displayed adenylate cyclase activity which was highly sensitive to forskolin and GPT analogues, whose action was antagonized by GDPβS. Receptor‐coupled regulation of adenylate cyclase thus appears to be mediated by mammalian‐type G proteins in C. elegans as well.
A consensus repeat sequence from the human insulin gene linked polymorphic region adopts multiple quadriplex DNA structures in vitro
Authors: Hammond-Kosack Michael C.U. , Docherty Keyin
Published: July 2002

A hypervariable region consisting of repeats of a 14 base pair (bp) consensus sequence ACAGGGGT(G/C)(T/C)GGGG is located 363 bp upstream of the human insulin gene. Different repeat numbers if this oligonucleotide give rise to a polymorphism, and so this region is commonly known as the insulin gene linked polymorphic region (ILPR). Here we present evidence, based on the mobility in non‐denaturing polyacrylamide gels of two dissimilarly sized oligonucleotides containing the ILPR consensus sequence, that this sequence can adopt a number of quadriplex DNA structures in vitro.
Rapid kinetics of the interaction between daunomycin and drug‐sensitive or drug‐resistant P388 leukemia cells
Authors: Soto Florentina , Canaves Jaume M. , Gonzalez-Ros Jose M. , Ferragut Jose A.
Published: July 2002

The initial stages of the interaction of daunomycin (DNM) with drug‐sensitive (P388/S) and drug‐resistant (P388/100) cells have been defined by a rapid kinetics stopped‐flow procedure. The process can be described by two kinetic components. The faster component accounts for rapid occupation of cell surface sites by DNM, as supported by experiments with liposomes with different surface charge. On the other hand, the effect of verapamil in the assays, suggests that the slower component is involved in the transport of the drug into the cells. Our observations are consistent with a loss in the control of the passive permeability to the drugs in the drug‐resistant tumor cells.
Truncation of recombinant vimentin by ompT Identification of a short motif in the head domain necessary for assembly of type III intermediate filament proteins
Authors: Hatzfeld Mechthild , Dodemont Huub , Plessmann Uwe , Weber Klaus
Published: July 2002

Recombinant vimentin expressed in E. coli JM 101 cells is cleaved after cell lysis between arginines 11 and 12. The truncated vimentin is assembly incompetent. Expression of the same cDNA construct in BL21 cells, which lack the protease ompT, provides intact and polymerization‐competent vimentin. The ompT cleavage site is contained in a short sequence motif (YRRMF) shared by the head domains of type III and IV intermediate filament (IF) proteins. We propose that a related motif present in the N‐terminal 32 residues of λCII acounts for the known IF formation of a fusion protein formed with a truncated GFAP.
Parathyroid hormone activation of stretch‐activated cation channels in osteosarcoma cells (UMR‐106.01)
Authors: Duncan Randall L. , Hruska Keith A. , Misler Stanley
Published: July 2002

Cell‐attached patches of membrane of osteoblast‐like cells UMR‐106.01 respond to bath application of parathyroid hormone (PTH) with an increase in the average activity, as well as the single channel conductance, of a stretch‐activated non‐selective cation channel. Correlations with whole cell membrane potential and conductance changes are considered.
Expression of the ras‐related rab3a gene in insulinoma‐derived cell lines
Authors: Lankat-Buttgereit Brigitte , Göke Rüdiger , Fehmann Hans-C. , Niess Constanze , Göke Burkhard
Published: July 2002

This study was designed to search for the expression of the small‐molecular‐weight GTP‐binding protein rab3a in endocrine pancreatic cell lines. Total RNA was isolated from five different cell lines (RINm5F, RIN 104836, β‐TC1, HIT‐15, and INRI‐G9) and from whole rat brain. The expression of rab3a was analyzed by Northern blots. Similar as in brain two transcripts of 1300 and 1800 bp were detected in RIN‐cells at low stringency conditions with the predominant signal at 1300 bp. At high stringency the stronger signal was at 1800 bp. When a 300 bp PstI fragment derived from the coding region of rab3a was utilized as probe the 1800 bp signal was predominant under each condition. Only a faint band at 1800 bp occurred in preparations from βTCI‐cells and no signal at all was found in HIT‐15 and INRI‐G9‐cells. In conclusion, rab3a is expressed in rat insulin‐releasing insulinoma‐derived RIN‐cells with a specific 1800 bp transcription product.
The levels of lactase and of sucrase‐isomaltase along the rabbit small intestine are regulated both at the mRNA level and post‐translationally
Authors: Keller Patrick , Zwicker Elmar , Mantei Ned , Semenza Giorgio
Published: July 2002

We determined along the small intestine of young and adult rabbits the activities of lactase (LPH) and sucrase (SI), the levels of their cognate mRNAs, and examined the in vitro biosynthesis of LPH and pro‐SI. Lactase activity is low in the proximal 1/3 of the intestine, whereas the mRNA levels are high. However, the rates of biosynthesis of the LPH forms correlated well with the steady‐state levels of LPH mRNA in all segments, indicating that factor(s) acting post‐translationally produce a decline in brush border LPH in the proximal small intestine. These factor(s) are not involved in the processing of pro‐LPH to mature LPH, since the relative amounts of the various forms of LPH are almost the same along the small intestine. Unexpectedly, we find that also for SI the ratio of activity to mRNA is low in proximal intestine. The biosynthesis of pro‐SI correlates with the steady‐state levels of its mRNA. Hence, the steady‐state levels of LPH and SI along the small intestine are regulated both by mRNA levels and by posttranslational factor(s).
Evidence for an interaction between cytosolic aldolase and the ATP‐ and pyrophosphate‐dependent phosphofructokinases in carrot storage roots
Authors: Moorhead Greg B.G. , Plaxton William C.
Published: July 2002

Immunoaffinity chromatography was employed to identify potential plant cytosolic aldolase (ALDc) binding proteins. A clarified homogenate of carrot storage root was chromatographed on a column of protein‐A—Sepharose that had been covalently coupled to anti‐(carrot root ALD,) immunoglobulin G. The column was washed with phosphate‐buffered saline (PBS), followed by step‐wise elution with increasing concentrations of NaCl in PBS. Several proteins were eluted following application of the salt gradient. Western blotting identified the major eluting proteins to be the PPi‐dependent phosphofructokinase (PFP) and the cytosolic form of the ATP‐dependent phosphofructokinase (PFKc), enzymes that are metabolically sequential to ALDc. The results suggest that ALDc may specifically interact with PFP and PFKc in carrots.
Bone morphogenetic proteins (BMP‐2 and BMP‐3) induce the late phase expression of the proto‐oncogene c‐fos in murine osteoblastic MC3T3‐E1 cells
Authors: Ohta Shuichi , Hiraki Yuji , Shigeno Chohei , Suzuki Fujio , Kasai Ryuichi , Ikeda Toshihiko , Kohno Hiroaki , Lee Kaechoong , Kikuchi Haruki , Konishi Junji , Bentz Hanne , Rosen David M. , Yamamuro Takao
Published: July 2002

Here we report that bone morphogenetic proteins 2 and 3 (BMP‐2 and BMP‐3) induced marked expression of c‐fos mRNA in a biphasic manner, i.e. the late phase (48 to 60 h) as well as the immediate‐early phase (0.5 h), in murine osteoblastic MC3T3‐El cells in vitro. The BMP‐induced late phase c‐fos gene expression was temporally associated with the onset of marked expression of the genes for osteocalcin and alkaline phosphatase, differentiation markers of mature osteoblasts. In contrast, none of TGF‐β1, 10% FBS, IGF‐I and IGF‐II, which induced only the immediate‐early c‐fos mRNA expression, stimulated the expression of osteocalcin and alkaline phosphatase genes. These data suggest that in osteoblasts BMP‐2 and BMP‐3 induce the late phase expression of c‐fos, which may play a role in transcriptional activation of the genes involved in differentiation of osteoblasts.
The thermal stability of the tryptic fragment of bovine microsomal cytochrome b5 and a variant containing six additional residues
Authors: Newbold Richard J. , Hewson Roger , Whitford David
Published: July 2002

Thermally induced denaturation has been measured for both oxidised and reduced forms of the tryptic fragment or bovine microsomal cytochrome b
5 using spectrophotometric methods. In the oxidised state, the tryptic fragment of cytochrome b
5 (Ala7‐Lys90) denatures in a single cooperative transition with a midpoint temperature (T
m) of ∼ 67°C (pH 7.0). The reduced form of the tryptic fragment of cytochrome b
5 shows a higher transition temperature of ∼ 73°C at pH 7.0 and this is reflected in the values of ΔH
m, ΔS
m, and Δ(ΔG) of ∼ 310kJ · mol−1, 900J · mol−1 · K−1 and 5 kJ · mol−1. Increased thermal stability is demonstrated for a variant protein that contains the first 90 amino acid residues of cytochrome b
5. These novel increases in stability are observed in both redox states and result from the presence of six additional residues at the amino‐terminus. The two forms of cytochrome b
5 do not differ significantly in structure with the results suggesting that the reorganisation energy (λ) of the variant protein, as measured indirectly from redox‐linked differences in conformational stability, is small. Consequently the reported subtle differences in reactivity between variants of cytochrome b
5 may result from the presence of additional N‐terminal residues on the surface of the protein.
Chronic ethanol administration enhances retinoic acid and triiodothyronine receptor expression in mouse liver
Authors: Pallet V. , Coustaut M. , Naulet F. , Higueret D. , Garcin H. , Higueret P.
Published: July 2002

Chronic alcoholism induces perturbations of storage and metabolization of retinol and related compounds. After 6 months of ethanol consumption we have observed in mouse liver an increased expression of Tri‐iodothyronine receptors (TR) while the expression of retinoic acid (RA) receptors (RAR) was unaffected. After 10 months of alcoholization the TR expression was strongly increased and the RAR expression was also increased. At this time the activity of aldehyde dehydrogenase and that of alcohol dehydrogenase, two enzymes involved in biosynthesis of RA from retinol, were similar in the liver of alcoholized and pair‐fed mice. Thus it can be hypothesized that (i) the change of RAR expression was, at least in part, the result of a change of TR expression (result in agreement with previous data), (ii) the increased expression of RAR could induce apoptosis and subsequently liver necrosis.
Assignment of NH resonances in nucleic acids using natural abundance 15N‐1H correlation spectroscopy with spin‐echo and gradient pulses
Authors: Szewczak Alexander A. , Kellogg Gregory W. , Moore Peter B.
Published: July 2002

It is well known that 2D 15N‐1H correlation spectra can resolve overlapped imino proton resonances in the downfleld NMR spectra of nucleic acids according to their 15N chemical shifts, and that these resonances can be assigned by base type on that basis, independent of conformation. This information can be extremely important in determining the solution structure of a nucleic acid by NMR, but previously could only be obtained using 15N‐labeled, or very concentrated samples. Here we report the design of a gradient‐enhanced, jump‐return spin echo version of an 15N‐1H HMQC experiment (GE‐JRSE HMQC) that is sensitive enough to work on unlabeled nucleic acid samples at normal NMR concentrations. This experiment has led to the assignment of imino proton resonances with non‐Watson‐Crick chemical shifts in the spectrum of a 29 residue oligoribonucleotide that models the sarcin/ricin loop from 28S rRNA.
Immunochemical identification of exposed regions of the Na+,K+‐ATPase α‐subunit
Authors: Homareda Haruo , Nagano Yoshine , Matsui Hideo
Published: July 2002

A polyclonal antibody against the Na+,K+‐ATPase holoenzyme was prepared. This antibody recognized native Na+,K+‐ATPase and inhibited its activity. The peptide fragments corresponding to various regions of the Na+,K+‐ATPase α‐subunit, which were synthesized from the cDNA, were immunoprecipitated with the antibody, and the M32‐D75, M158‐D197 and M470‐V552 fragments (the latter included K508, a putative ATP binding site) were identified as the epitopes. The M267‐I442 fragment, which included a phosphorylation site at D376, and the C‐terminal one‐third of the α‐subunit from M615 to the C‐terminus, were not detected using this antibody. These results suggest that at least three regions on the Na+,K+‐ATPase α‐subunit, M32‐D75, M158‐D197 and M470‐V552, cover its exposed regions, and that some of them are essential for ATPase activity.
Molecular cloning of two receptors from rat brain with high affinity for salmon calcitonin
Authors: Albrandt Keith , Mull Emily , Brady Elize M.G. , Herich John , Moore Candace X. , Beaumont Kevin
Published: July 2002

Two receptors with high affinity for salmon calcitonin were cloned from the nucleus accumbens region of rat brain. The deduced 479 amino acid sequence of cDNA clone L2175‐D20 (designated Cla receptor) is 78% and 66% identical with those reported for human and porcine calcitonin receptors, respectively. Clone U3237‐A2 codes for a receptor (designated Clb) that is identical to Cla except for a 37 amino acid insert in the second extracellular domain. COS‐7 cells transfected with either transcript bound [125I]salmon calcitonin with high affinity (K
d = 8 pM for Cla; K
d = 48 pM for Clb) and responded to salmon calcitonin with increases in cAMP. Tissue distribution studies revealed Cla transcript in rat brain, skeletal muscle, kidney and lung, whereas Clb was predominantly found in brain.
Enhancing factor protein from mouse small intestines belongs to the phospholipase A2 family
Authors: Mulherkar Rita , Rao Renuka , Rao Laxmi , Patki Varsha , Chauhan V.S. , Deo Madhav G.
Published: July 2002

Enhancing factor (EF), a growth regulatory molecule, isolated from mouse small intestines, has been well characterized in this laboratory. It increases the binding of epidermal growth factor in a unique manner via its own receptor. In the first 20 N‐terminal amino acids sequenced, EF showed 50% homology to human Group II phospholipase A2 (PLA2). Here we propose that EF is yet another, unidentified isoform ofPLA2 which regulates cell proliferation via modulation of EGF binding. To our knowledge, this is the first report implicating PLA2‐II‐like molecules in growth regulation.
P1, P3‐bis(5′‐adenosyl)triphosphate (Ap3A) as a substrate and a product of mammalian tryptophanyl‐tRNA synthetase
Authors: Merkulova Tatyana , Kovaleva Galina , Kisselev Lev
Published: July 2002

Bovine tryptophanyl‐tRNA synthetase (TrpRS, E.C. 6.1.1.2) is unable to catalyze in vitro formation of Ap4A in contrast to some other aminoacyl‐tRNA synthetases. However, in the presence of l‐tryptophan, ATP‐Mg2+ and ADP the enzyme catalyzes the Ap3A synthesis via adenylate intermediate. Ap3A (not Ap4A) may serve as a substrate for TrpRS in the reaction of E·(Trp ∼ AMP) formation and in the tRNATrp charging. The K
m value for Ap3A was higher than the K
m for ATP (approx. 1.00 vs. 0.22 mM) and V
max was 3 times lower than for ATP. The Zn2+‐deficient enzyme catalyzes Ap3A synthesis in the absence of exogenous ADP due to ATPase activity of Zn2+‐deprived TrpRS. The inability of mammalian TrpRS to synthesize Ap4A, might be considered as a molecular tool preventing the removal of Zn2+ due to chelation by Ap4A and therefore preserving the enzyme activity.
DNA binding activity of NtrC from Rhizobium grown on different nitrogen sources
Authors: Patriarca Eduardo J. , Riccio Anna , Colonna-Romano Sergio , Defez Roberto , Iaccarino Maurizio
Published: July 2002

The DNA‐binding activity of the NtrC protein can be demonstrated in gel retardation assays with concentrated protein extracts of Rhizobium etli. Using extracts from either the wild type or a ntrC mutant strain and an antiserum raised against the NtrC protein, we demonstrate specific binding of NtrC to the upstream regulatory region of the glnII gene, where two putative NtrC‐binding sites are present. KNO3‐grown bacteria contain less NtrC protein and more NtrC‐binding activity than NH4Cl‐grown bacteria, thus showing that with this protocol it is possible to detect changes in NtrC‐binding activity. The advantages of this assay system in comparison with that using pure proteins is discussed.