bluuebunny/crossref_metadata_2025_split huggingface dataset:
Stress-Induced Legume Root Nodule Senescence. Physiological, Biochemical, and Structural Alterations
Authors: Matamoros Manuel A. , Baird Lisa M. , Escuredo Pedro R. , Dalton David A. , Minchin Frank R. , Iturbe-Ormaetxe Iñaki , Rubio Maria C. , Moran Jose F. , Gordon Anthony J. , Becana Manuel 
Published: July 2002

AbstractNitrate-fed and dark-stressed bean (Phaseolus vulgaris) and pea (Pisum sativum) plants were used to study nodule senescence. In bean, 1 d of nitrate treatment caused a partially reversible decline in nitrogenase activity and an increase in O2 diffusion resistance, but minimal changes in carbon metabolites, antioxidants, and other biochemical parameters, indicating that the initial decrease in nitrogenase activity was due to O2 limitation. In pea, 1 d of dark treatment led to a 96% decline in nitrogenase activity and sucrose, indicating sugar deprivation as the primary cause of activity loss. In later stages of senescence (4 d of nitrate or 2–4 d of dark treatment), nodules showed accumulation of oxidized proteins and general ultrastructural deterioration. The major thiol tripeptides of untreated nodules were homoglutathione (72%) in bean and glutathione (89%) in pea. These predominant thiols declined by approximately 93% after 4 d of nitrate or dark treatment, but the loss of thiol content can be only ascribed in part to limited synthesis by γ-glutamylcysteinyl, homoglutathione, and glutathione synthetases. Ascorbate peroxidase was immunolocalized primarily in the infected and parenchyma (inner cortex) nodule cells, with large decreases in senescent tissue. Ferritin was almost undetectable in untreated bean nodules, but accumulated in the plastids and amyloplasts of uninfected interstitial and parenchyma cells following 2 or 4 d of nitrate treatment, probably as a response to oxidative stress.
Antisense HEMA1 RNA Expression Inhibits Heme and Chlorophyll Biosynthesis in Arabidopsis
Authors: Kumar A. Madan , Söll Dieter 
Published: July 2002

Abstract
5-Aminolevulinic acid (ALA) is a precursor in the biosynthesis of tetrapyrroles including chlorophylls and heme. The formation of ALA involves two enzymatic steps which take place in the chloroplast in plants. The first enzyme, glutamyl-tRNA reductase, and the second enzyme, glutamate-1-semialdehyde-2,1-aminomutase, are encoded by the nuclearHEMA and GSA genes, respectively. To assess the significance of the HEMA gene for chlorophyll and heme synthesis, transgenic Arabidopsis plants that expressed antisense HEMA1 mRNA from the constitutive cauliflower mosaic virus 35S promoter were generated. These plants exhibited varying degrees of chlorophyll deficiency, ranging from patchy yellow to total yellow. Analysis indicated that these plants had decreased levels of chlorophyll, non-covalently bound hemes, and ALA; their levels were proportional to the level of glutamyl-tRNA reductase expression and were inversely related to the levels of antisenseHEMA transcripts. Plants that lacked chlorophyll failed to survive under normal growth conditions, indicating thatHEMA gene expression is essential for growth.
CHRK1, a Chitinase-Related Receptor-Like Kinase in Tobacco
Authors: Kim Youn Sung , Lee Jeong Hee , Yoon Gyeong Mee , Cho Hye Sun , Park Seong-Whan , Suh Mi Chung , Choi Doil , Ha Hyun Jung , Liu Jang Ryol , Pai Hyun-Sook 
Published: July 2002

AbstractA cDNA encoding a chitinase-related receptor-like kinase, designated CHRK1, was isolated from tobacco (Nicotiana tabacum). The C-terminal kinase domain (KD) of CHRK1 contained all of the conserved amino acids of serine/threonine protein kinases. The putative extracellular domain was closely related to the class V chitinase of tobacco and to microbial chitinases.CHRK1 mRNA accumulation was strongly stimulated by infection with fungal pathogen and tobacco mosaic virus. Amino acid-sequence analysis revealed that the chitinase-like domain of CHRK1 lacked the essential glutamic acid residue required for chitinase activity. The recombinant chitinase-like domain did not show any catalytic activity for either oligomeric or polymeric chitin substrates. The recombinant KD of CHRK1 exhibited autophosphorylation, but the mutant KD with a mutation in the essential ATP-binding site did not, suggesting that CHRK1 encoded a functional kinase. CHRK1 was detected in membrane fractions of tobacco BY2 cells. Furthermore, CHRK1-GFP fusion protein was localized in plasma membranes when it was expressed in animal cells. This is the first report of a new type of receptor-like kinase containing a chitinase-like sequence in the putative extracellular domain.
Dehydration-Induced Redistribution of Amphiphilic Molecules between Cytoplasm and Lipids Is Associated with Desiccation Tolerance in Seeds
Authors: Buitink Julia , Leprince Olivier , Hoekstra Folkert A. 
Published: July 2002

AbstractThis study establishes a relationship between desiccation tolerance and the transfer of amphiphilic molecules from the cytoplasm into lipids during drying, using electron paramagnetic resonance spectroscopy of amphiphilic spin probes introduced into imbibed radicles of pea (Pisum sativum) and cucumber (Cucumis sativa) seeds. Survival following drying and a membrane integrity assay indicated that desiccation tolerance was present during early imbibition and lost in germinated radicles. In germinated cucumber radicles, desiccation tolerance could be re-induced by an incubation in polyethylene glycol (PEG) before drying. In desiccation-intolerant radicles, partitioning of spin probes into lipids during dehydration occurred at higher water contents compared with tolerant and PEG-induced tolerant radicles. The difference in partitioning behavior between desiccation-tolerant and -intolerant tissues could not be explained by the loss of water. Consequently, using a two-phase model system composed of sunflower or cucumber oil and water, physical properties of the aqueous solvent that may affect the partitioning of amphiphilic spin probes were investigated. A significant relationship was found between the partitioning of spin probes and the viscosity of the aqueous solvent. Moreover, in desiccation-sensitive radicles, the rise in cellular microviscosity during drying commenced at higher water contents compared with tolerant or PEG-induced tolerant radicles, suggesting that the microviscosity of the cytoplasm may control the partitioning behavior in dehydrating seeds.
Chromosaponin I Specifically Interacts with AUX1 Protein in Regulating the Gravitropic Response of Arabidopsis Roots
Authors: Rahman Abidur , Ahamed Arifa , Amakawa Taisaku , Goto Nobuharu , Tsurumi Seiji 
Published: July 2002

Abstract
We have found that chromosaponin I (CSI), a γ-pyronyl-triterpenoid saponin isolated from pea (Pisum sativum L. cv Alaska), specifically interacts with AUX1 protein in regulating the gravitropic response of Arabidopsis roots. Application of 60 μm CSI disrupts the vertically oriented elongation of wild-type roots grown on agar plates but orients the elongation of agravitropic mutant aux1-7 roots toward the gravity. The CSI-induced restoration of gravitropic response inaux1-7 roots was not observed in other agravitropic mutants, axr2 and eir1-1. Because theaux1-7 mutant is reduced in sensitivity to auxin and ethylene, we examined the effects of CSI on another auxin-resistant mutant, axr1-3, and ethylene-insensitive mutantein2-1. In aux1-7 roots, CSI stimulated the uptake of [3H]indole-3-acetic acid (IAA) and induced gravitropic bending. In contrast, in wild-type, axr1-3, and ein2-1 roots, CSI slowed down the rates of gravitropic bending and inhibited IAA uptake. In the null allele ofaux1, aux1-22, the agravitropic nature of the roots and IAA uptake were not affected by CSI. This close correlation between auxin uptake and gravitropic bending suggests that CSI may regulate gravitropic response by inhibiting or stimulating the uptake of endogenous auxin in root cells. CSI exhibits selective influence toward IAA versus 1-naphthaleneacetic acid as to auxin-induced inhibition in root growth and auxin uptake. The selective action of CSI toward IAA along with the complete insensitivity of the null mutant aux1-22 toward CSI strongly suggest that CSI specifically interacts with AUX1 protein.
Solution structure of a yeast ubiquitin‐like protein Smt3: The role of structurally less defined sequences in protein–protein recognitions
Authors: Sheng Wanyun , Liao Xiubei 
Published: July 2002

AbstractSmt3 belongs to a growing family of ubiquitin‐related proteins involved in posttranslational protein modification. Independent studies demonstrate an essential function of Smt3 in the regulation of nucleocytoplasmic transport, and suggest a role in cell‐cycle regulation. Here we report the high‐resolution NMR structure of yeast Smt3 in the complex free form. Our comparison of the Smt3 NMR structure with the Smt3 crystal structure in complex with the C‐Terminal Ulp1 protease domain revealed large structural differences in the binding surface, which is also involved in the Smt3‐Ubc‐9 interaction detected by NMR. The structural differences in the region indicate the important functions of conserved residues in less structurally defined sequences.
Threading of a glycosylated protein loop through a protein hole: Implications for combination of human chorionic gonadotropin subunits
Authors: Xing Yongna , Williams Christopher , Campbell Robert K. , Cook Stephanie , Knoppers Maria , Addona Theresa , Altarocca Valter , Moyle William R. 
Published: July 2002

AbstractChorionic gonadotropin (hCG) is a heterodimeric placental glycoprotein hormone essential for human reproduction. Twenty hCG β‐subunit residues, termed the seatbelt, are wrapped around α‐subunit loop 2 (α2) and their positions “latched” by a disulfide formed by cysteines at the end of the seatbelt (Cys 110) and in the β‐subunit core (Cys 26). This unique arrangement explains the stability of the heterodimer but raises questions as to how the two subunits combine. The seatbelt is latched in the free β‐subunit. If the seatbelt remained latched during the process of subunit combination, formation of the heterodimer would require α2 and its attached oligosaccharide to be threaded through a small β‐subunit hole. The subunits are known to combine during oxidizing conditions in vitro, and studies described here tested the idea that this requires transient disruption of the latch disulfide, possibly as a consequence of the thioredoxin activity reported in hCG. We observed that alkylating agents did not modify either cysteine in the latch disulfide (Cys 26 or Cys 110) during heterodimer formation in several oxidizing conditions and had minimal influence on these cysteines during combination in the presence of mild reductants (1–3 mM β‐mercaptoethanol). Reducing agents appeared to accelerate subunit combination by disrupting a disulfide (Cys 93–Cys 100) that forms a loop within the seatbelt, thereby increasing the size of the β‐subunit hole. We propose a mechanism for hCG assembly in vitro that depends on movements of α2 and the seatbelt and suggest that the process of glycoprotein hormone subunit combination may be useful for studying the movements of loops during protein folding.
An extended hudrophobic core induces EF‐hand swapping
Authors: Håkansson Maria , Svensson Anders , Fast Jonas , Linse Sara 
Published: July 2002

AbstractThe structure of calbindin D9k with two substitutions was determined by X‐ray crystallography at 1.8‐Å resolution. Unlike wild‐type calbindin D9k, which is a monomeric protein with two EF‐hands, the structure of the mutated calbindin D9k reveals an intertwined dimer. In the dimer, two EF‐hands of the monomers have exchanged places, and thus a 3D domain‐swapped dimer has been formed. EF‐hand I of molecule A is packed toward EF‐hand II of molecule B and vice versa. The formation of a hydrophobic cluster, in a region linking the EF‐hands, promotes the conversion of monomers to 3D domain‐swapped dimers. We propose a mechanism by which domain swapping takes place via the apo form of calbindin D9k. Once formed, the calbindin D9k dimers are remarkably stable, as with even larger misfolded aggregates like amyloids. Thus calbindin D9k dimers cannot be converted to monomers by dilution. However, heating can be used for conversion, indicating high energy barriers separating monomers from dimers.
NMR and SAXS characterization of the denatured state of the chemotactic protein Che Y: Implications for protein folding initiation
Authors: Garcia Pascal , Serrano Luis , Durand Dominique , Rico Manuel , Bruix Marta 
Published: July 2002

AbstractThe denatured state of a double mutant of the chemotactic protein CheY (F14N/V83T) has been analyzed in the presence of 5 M urea, using small angle X‐ray scattering (SAXS) and heteronuclear magnetic resonance. SAXS studies show that the denatured protein follows a wormlike chain model. Its backbone can be described as a chain composed of rigid elements connected by flexible links. A comparison of the contour length obtained for the chain at 5 M urea with the one expected for a fully expanded chain suggests that ∼25% of the residues are involved in residual structures. Conformational shifts of the α‐protons, heteronuclear 15N‐{1H} NOEs and 15N relaxation properties have been used to identify some regions in the protein that deviate from a random coil behavior. According to these NMR data, the protein can be divided into two subdomains, which largely coincide with the two folding subunits identified in a previous kinetic study of the folding of the protein. The first of these subdomains, spanning residues 1–70, is shown here to exhibit a restricted mobility as compared to the rest of the protein. Two regions, one in each subdomain, were identified as deviating from the random coil chemical shifts. Peptides corresponding to these sequences were characterized by NMR and their backbone 1H chemical shifts were compared to those in the intact protein under identical denaturing conditions. For the region located in the first subdomain, this comparison shows that the observed deviation from random coil parameters is caused by interactions with the rest of the molecule. The restricted flexibility of the first subdomain and the transient collapse detected in that subunit are consistent with the conclusions obtained by applying the protein engineering method to the characterization of the folding reaction transition state.
Mass spectrometric study of the Escherichia coli repressor proteins, IcIR and GcIR, and their complexes with DNA
Authors: Donald Lynda J. , Hosfield David J. , Cuvelier Susan L. , Ens Werner , Standing Kenneth G. , Duckworth Harry W. 
Published: July 2002

AbstractIn Escherichia coli, the IclR protein regulates both the aceBAK operon and its own synthesis. Database homology searches have identified many IclR‐like proteins, now known as the IclR family, which can be identified by a conserved C‐terminal region. We have cloned and purified one of these proteins, which we have named GclR (glyoxylate carboligase repressor). Although purification is straightforward, both the IclR and GclR proteins are difficult to manipulate, requiring high salt (up to 0.6 M KCl) for solubility. With the advent of nanospray ionization, we could transfer the proteins into much higher concentrations of volatile buffer than had been practical with ordinary electrospray. In 0.5 M ammonium bicarbonate buffer, both proteins were stable as tetramers, with a small amount of dimer. In a separate experiment, we found that IclR protein selected from a random pool a sequence which matched exactly that of the presumed binding region of the GclR protein, although IclR does not regulate the gcl gene. We designed a 29 bp synthetic DNA to which IclR and GclR bind, and with which we were able to form noncovalent DNA‐protein complexes for further mass spectrometry analysis. These complexes were far more stable than the proteins alone, and we have evidence of a stoichiometry which has not been described previously with (protein monomer : dsDNA) = (4 : 1).
Computational Design and Experimental Evaluation of Using a Leading Edge Fillet on a Gas Turbine Vane
Authors: Zess G. A. , Thole K. A. 
Published: July 2002

AbstractWith the desire for increased power output for a gas turbine engine comes the continual push to achieve higher turbine inlet temperatures. Higher temperatures result in large thermal and mechanical stresses particularly along the nozzle guide vane. One critical region along a vane is the leading edge-endwall juncture. Based on the assumption that the approaching flow to this juncture is similar to a two-dimensional boundary layer, previous studies have shown that a horseshoe vortex forms. This vortex forms because of a radial total pressure gradient from the approaching boundary layer. This paper documents the computational design and experimental validation of a fillet placed at the leading edge-endwall juncture of a guide vane to eliminate the horseshoe vortex. The fillet design effectively accelerated the incoming boundary layer thereby mitigating the effect of the total pressure gradient. To verify the CFD studies used to design the leading edge fillet, flowfield measurements were performed in a large-scale, linear, vane cascade. The flowfield measurements were performed with a laser Doppler velocimeter in four planes orientated orthogonal to the vane. Good agreement between the CFD predictions and the experimental measurements verified the effectiveness of the leading edge fillet at eliminating the horseshoe vortex. The flow-field results showed that the turbulent kinetic energy levels were significantly reduced in the endwall region because of the absence of the unsteady horseshoe vortex.
Abstract: Ion implantation and proton−enhanced diffusion in semiconductors
Authors: Gibbons James F. 
Published: July 2002

In a period of less than ten years, ion implantation has become an accepted production process in the semiconductor industry, despite considerable initial skepticism about the merits of the process. More recently, proton−enhanced diffusion has been used to fabricate state−of−the−art microwave bipolar transistors and appears to have a number of other applications, including extensions of the frequency response of GaAs FETs.
The purpose of this paper is to describe the basic physics involved in each of these processes and to show how the relative merits of each lead naturally to a set of applications in which ion implantation, enhanced diffusion, and conventional thermal processes are combined to produce devices with characteristics superior to those previously available.
The principal topics to be presented are the following: (1) Basic operating features of implantation machines with special attention to those characteristics which are required for economic, efficient production; (2) Basic physics of the implantation process with special attention to range distribution theory and experiments, and damage production and annealing; (3) Applications of ion implantation to the production of semiconductor devices including MOSFETs, bipolar transistors, varactors, CCDs, and high−efficiency LEDs; and (4) Basic physics of the proton enhanced diffusion process and its emerging applications in the fabrication of microwave devices.
Abstract: Backscattering spectrometry
Authors: Mayer J. W. , Nicolet M−A. , Chu W. K. 
Published: July 2002

Backscattering spectrometry is the microanalysis of the surface and near−surface regions of materials utilizing energetic (typically 1−2 MeV) He and H ions as the analytical probe. 1−4 The ions in the collimated beam provided by a particle accelerator penetrate into the target and are scattered at various depths. Analysis of the energy spectrum of the particles backscattered from the target provides information in three areas: identification of atomic mass of the target constituents, the target composition, and depth profiles. Mass identification is provided by the kinematics of the energy transfer between the projectiles and target atom in the collision process. For MeV He ions, the mass resolution is ±1 atomic mass unit (amu) up to about mass 40, and deteriorates to about ±10 amu for the heaviest elements. Quantitative analysis of the number of target atoms per cm2 is provided by the scattering cross section which connects the measured yield of backscattered particles to target composition. In compositional analysis, backscattering only provides atomic ratios, for information on the chemical nature of the material, x−ray diffraction or other tools with chemical sensitivity must be used. Backscattering techniques are most useful to examine heavy elements in a light−element matrix as the sensitivity is rather poor for light elements unless proton resonance techniques are used.
Depth information is provided by the energy loss of the projectiles on their inward and outward paths through the sample. The energy loss of the projectiles per unit path length depends on the energy of the particle as well as on the projectile and target. Compilations (5) and reviews of stopping cross sections are available. For MeV 4He ions, the depth resolution is about 200−300 Å for depths up to 0.5 μ. At greater depths, energy straggling causes a degradation in depth resolution.
Surveys of the applications of backscattering spectrometry are provided in Ref. 1−4. The technique has been used to analyze ion−implanted samples, thin−deposited films, metal−metal thin film interdiffusion, metal film−bulk silicon reactions, solubility of impurities in bulk samples, oxidation processes and other phenomena occurring in the near−surface region of materials.
A typical beam spot is 1 mm2 in area, but for convenient handling, samples should preferably have a size of 1/2−1 cm2. The uniformity of the sample over the area of the beam spot is critical. Since backscattering analysis has no laterial resolution, the samples have to be checked with scanning electron microscopes or with optical microscpoes to verify this assumption.
The primary strength of backscattering analysis is the ability to provide quantitative information on atomic composition ratios and on absolute thicknesses of films in the range of several thousand Angstroms without resorting to standards. Routine accuracies for this information are of the order of ±5%.