bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of a Granule-Bound Starch Synthase Isoform Found in the Pericarp of Wheat
Authors: Nakamura Toshiki , Vrinten Patricia , Hayakawa Kazuhiro , Ikeda Junichi 
Published: July 2002

AbstractWaxy wheat (Triticum aestivum L.) lacks the waxy protein, which is also known as granule-bound starch synthase I (GBSSI). The starch granules of waxy wheat endosperm and pollen do not contain amylose and therefore stain red-brown with iodine. However, we observed that starch from pericarp tissue of waxy wheat stained blue-black and contained amylose. Significantly higher starch synthase activity was detected in pericarp starch granules than in endosperm starch granules. A granule-bound protein that differed from GBSSI in molecular mass and isoelectric point was detected in the pericarp starch granules but not in granules from endosperm. This protein was designated GBSSII. The N-terminal amino acid sequence of GBSSII, although not identical to wheat GBSSI, showed strong homology to waxy proteins or GBSSIs of cereals and potato, and contained the motif KTGGL, which is the putative substrate-binding site of GBSSI of plants and of glycogen synthase ofEscherichia coli. GBSSII cross-reacted specifically with antisera raised against potato and maize GBSSI. This study indicates that GBSSI and GBSSII are expressed in a tissue-specific manner in different organs, with GBSSII having an important function in amylose synthesis in the pericarp.
Quantitative Control of Inflorescence Formation inImpatiens balsamina
Authors: Pouteau Sylvie , Tooke Fiona , Battey Nicholas 
Published: July 2002

Abstract
We analyzed the process of inflorescence formation in Impatiens balsamina by studying the architecture of the plant under different photoperiod treatments. Floral reversion under noninductive conditions in this species is caused by the lack of persistence of the induced state in the leaf. This can be used to control the amount of inductive signal and to examine its quantitative influence on morphological changes in the plant. The floral transition was characterized by a continuum of variation at the level of meristem identity, primordium initiation, and floral organ identity. This continuum was enhanced during reversion, suggesting that the establishment of a continuum partly reflects limiting amounts of inductive signal exported from the leaf to the meristem. The transcription patterns of two homologs of genes involved in the control of floral meristem identity, Imp-FLO andImp-FIM, were similar in terminal and axillary flowers and may be associated with the continuum exhibited by I. balsamina. By analyzing the fate of axillary meristem primordia initiated before and after the beginning of the inductive period, we showed that de novo initiation of axillary meristem primordia by the evoked meristem is not required and that primordia initiated before evocation can adopt different fates, depending on the amount of inductive signal. The influence of age and/or position on primordium responsiveness to the inductive signal is discussed.
Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Content, Assimilatory Charge, and Mesophyll Conductance in Leaves1
Authors: Eichelmann Hillar , Laisk Agu 
Published: July 2002

Abstract
The content of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) (Et; EC 4.1.1.39) measured in different-aged leaves of sunflower (Helianthus annuus) and other plants grown under different light intensities, varied from 2 to 75 μmol active sites m−2. Mesophyll conductance (μ) was measured under 1.5% O2, as well as postillumination CO2uptake (assimilatory charge, a gas-exchange measure of the ribulose-1,5-bisphosphate pool). The dependence of μ onEt saturated atEt = 30 μmol active sites m−2and μ = 11 mm s−1 in high-light-grown leaves. In low-light-grown leaves the dependence tended toward saturation at similar Et but reached a μ of only 6 to 8 mm s−1. μ was proportional to the assimilatory charge, with the proportionality constant (specific carboxylation efficiency) between 0.04 and 0.075 μm−1s−1. Our data show that the saturation of the relationship between Et and μ is caused by three limiting components: (a) the physical diffusion resistance (a minor limitation), (b) less than full activation of Rubisco (related to Rubisco activase and the slower diffusibility of Rubisco at high protein concentrations in the stroma), and (c) chloroplast metabolites, especially 3-phosphoglyceric acid and free inorganic phosphate, which control the reaction kinetics of ribulose-1,5-bisphosphate carboxylation by competitive binding to active sites.
Dynamic Properties of Endogenous Phytochrome A in Arabidopsis Seedlings
Authors: Hennig Lars , Büche Claudia , Eichenberg Klaus , Schäfer Eberhard 
Published: July 2002

AbstractThe dynamic behavior of phytochrome A (phyA) in seedlings of the model plant Arabidopsis was examined by in vivo spectroscopy and by western and northern blotting. Rapid accumulation of phyA was observed, reaching a steady state after 3 d. Both red and far-red light initiated a rapid destruction of the far-red-light-absorbing form of phytochrome (Pfr); the apparent half-life was only 4-fold longer in far-red than in red light. Furthermore, the Pfr-induced destruction of the red-light-absorbing form of phytochrome (Pr) of phyA occurred in darkness with a rate identical to that of Pfr destruction. A 2-fold decrease in mRNA abundance was observed after irradiation, irrespective of the applied light quality. However, reaccumulation occurred rapidly after far-red but slowly after red irradiation, indicating different modes of regulation of phytochrome expression after light-dark transitions depending on the light quality of the preceding irradiation. The wavelength dependency of the destruction rates was distinct from that of mustard, a close relative of Arabidopsis, and was explained on the basis of Pfr-induced Pr destruction and a simple kinetic two-step model. No dark reversion was detectable in the destruction kinetics after a red pulse. From these data we conclude that Arabidopsis phyA differs significantly in several aspects from other dicot phytochromes.
The Allosterically Unregulated Isoform of ADP-Glucose Pyrophosphorylase from Barley Endosperm Is the Most Likely Source of ADP-Glucose Incorporated into Endosperm Starch
Authors: Doan Danny N.P. , Rudi Heidi , Olsen Odd-Arne 
Published: July 2002

Abstract
We present the results of studies of an unmodified version of the recombinant major barley (Hordeum vulgare) endosperm ADP-glucose pyrophoshorylase (AGPase) expressed in insect cells, which corroborate previous data that this isoform of the enzyme acts independently of the allosteric regulators 3-phosphoglycerate and inorganic phosphate. We also present a characterization of the individual subunits expressed separately in insect cells, showing that the SS AGPase is active in the presence of 3-phosphoglycerate and is inhibited by inorganic phosphate. As a step toward the elucidation of the role of the two AGPase isoforms in barley, the temporal and spatial expression profile of the four barley AGPase transcripts encoding these isoforms were studied. The results show that the steady-state level ofbeps and bepl, the transcripts encoding the major endosperm isoform, correlated positively with the rate of endosperm starch accumulation. In contrast, blps andblpl, the transcripts encoding the major leaf isoform, were constitutively expressed at a very low steady-state level throughout the barley plant. The implications of these findings for the evolution of plant AGPases are discussed.
Proline Metabolism in the Wild-Type and in a Salt-Tolerant Mutant of Nicotiana plumbaginifolia Studied by13C-Nuclear Magnetic Resonance Imaging
Authors: Roosens Nancy H. , Willem Rudolph , Li Yan , Verbruggen Ingrid , Biesemans Monique , Jacobs Michel 
Published: July 2002

Abstract
To obtain insight into the link between proline (Pro) accumulation and the increase in osmotolerance in higher plants, we investigated the biochemical basis for the NaCl tolerance of a Nicotiana plumbaginifolia mutant (RNa) that accumulates Pro. Pro biosynthesis and catabolism were investigated in both wild-type and mutant lines. 13C-Nuclear magnetic resonance with [5-13C]glutamate (Glu) as the Pro precursor was used to provide insight into the mechanism of Pro accumulation via the Glu pathway. After 24 h under 200 mm NaCl stress in the presence of [5-13C]Glu, a significant enrichment in [5-13C]Pro was observed compared with non-stress conditions in both the wild type (P2) and the mutant (RNa). Moreover, under the same conditions, [5-13C]Pro was clearly synthesized in higher amounts in RNa than in P2. On the other hand, measurements of enzyme activities indicate that neither the biosynthesis via the ornithine pathway, nor the catabolism via the Pro oxidation pathway were affected in the RNa mutant. Finally, the regulatory effect exerted by Pro on its biosynthesis was evaluated. In P2 plantlets, exogenous Pro markedly reduced the conversion of [5-13C]Glu into [5-13C]Pro, whereas Pro feedback inhibition was not detected in the RNa plantlets. It is proposed that the origin of tolerance in the RNa mutant is due to a mutation leading to a substantial reduction of the feedback inhibition normally exerted in a wild-type (P2) plant by Pro at the level of the Δ-pyrroline-5-carboxylate synthetase enzyme.
Amyloplast Sedimentation Dynamics in Maize Columella Cells Support a New Model for the Gravity-Sensing Apparatus of Roots
Authors: Yoder Thomas L. , Zheng Hui-qiong , Todd Paul , Staehelin L. Andrew 
Published: July 2002

AbstractQuantitative analysis of statolith sedimentation behavior was accomplished using videomicroscopy of living columella cells of corn (Zea mays) roots, which displayed no systematic cytoplasmic streaming. Following 90° rotation of the root, the statoliths moved downward along the distal wall and then spread out along the bottom with an average velocity of 1.7 μm min−1. When statolith trajectories traversed the complete width or length of the cell, they initially moved horizontally toward channel-initiation sites and then moved vertically through the channels to the lower side of the reoriented cell where they again dispersed. These statoliths exhibited a significantly lower average velocity than those sedimenting on distal-to-side trajectories. In addition, although statoliths undergoing distal-to-side sedimentation began at their highest velocity and slowed monotonically as they approached the lower cell membrane, statoliths crossing the cell's central region remained slow initially and accelerated to maximum speed once they reached a channel. The statoliths accelerated sooner, and the channeling effect was less pronounced in roots treated with cytochalasin D. Parallel ultrastructural studies of high-pressure frozen-freeze-substituted columella cells suggest that the low-resistance statolith pathway in the cell periphery corresponds to the sharp interface between the endoplasmic reticulum (ER)-rich cortical and the ER-devoid central region of these cells. The central region is also shown to contain an actin-based cytoskeletal network in which the individual, straight, actin-like filaments are randomly distributed. To explain these findings as well as the results of physical simulation experiments, we have formulated a new, tensegrity-based model of gravity sensing in columella cells. This model envisages the cytoplasm as pervaded by an actin-based cytoskeletal network that is denser in the ER-devoid central region than in the ER-rich cell cortex and is linked to stretch receptors in the plasma membrane. Sedimenting statoliths are postulated to produce a directional signal by locally disrupting the network and thereby altering the balance of forces acting on the receptors in different plasma membrane regions.
Homeodomain Leucine Zipper Proteins Bind to the Phosphate Response Domain of the SoybeanVspBTripartite Promoter
Authors: Tang Zhijun , Sadka Avi , Morishige Daryl T. , Mullet John E. 
Published: July 2002

AbstractThe soybean (Glycine max L. Merr. cv Williams 82) genes VspA and VspB encode vacuolar glycoprotein acid phosphatases that serve as vegetative storage proteins during seed fill and early stages of seedling growth.VspB expression is activated by jasmonates (JAs) and sugars and down-regulated by phosphate and auxin. Previous promoter studies demonstrated that VspB promoter sequences between −585 and −535 mediated responses to JA, and sequences between −535 and −401 mediated responses to sugars, phosphate, and auxin. In this study, the response domains were further delineated using transient expression of VspBpromoter-β-glucuronidase constructs in tobacco protoplasts. Sequences between −536 and −484 were identified as important for phosphate responses, whereas the region from −486 to −427 mediated sugar responses. Gel-shift and deoxyribonuclease-I footprinting assays revealed four DNA-binding sites between −611 and −451 of the soybeanVspB promoter: one in the JA response domain, two in the phosphate response domain, and one binding site in the sugar response domain. The sequence CATTAATTAG present in the phosphate response domain binds soybean homeodomain leucine zipper proteins, suggesting a role for these transcription factors in phosphate-modulated gene expression.
The Cyclization of Farnesyl Diphosphate and Nerolidyl Diphosphate by a Purified Recombinant δ-Cadinene Synthase
Authors: Benedict Chauncey R. , Lu Jia-Ling , Pettigrew Donald W. , Liu Jinggao , Stipanovic Robert D. , Williams Howard J. 
Published: July 2002

Abstract
The first step in the conversion of the isoprenoid intermediate, farnesyl diphosphate (FDP), to sesquiterpene phytoalexins in cotton (Gossypium barbadense) plants is catalyzed by δ-cadinene (CDN) synthase. CDN is the precursor of desoxyhemigossypol and hemigossypol defense sesquiterpenes. In this paper we have studied the mechanism for the cyclization of FDP and the putative intermediate, nerolidyl diphosphate, to CDN. A purified recombinant CDN synthase (CDN1-C1) expressed in Escherichia colifrom CDN1-C1 cDNA isolated from Gossypium arboreumcyclizes (1RS)-[1-2H](E, E)-FDP to &gt;98% [5-2H]and [11-2H]CDN. Enzyme reaction mixtures cyclize (3RS)-[4,4,13,13,13-2H5]-nerolidyl diphosphate to 62.1% [8,8,15,15,15-2H5]-CDN, 15.8% [6,6,15,15,15-2H5]-α-bisabolol, 8.1% [6,6,15,15,15-2H5]-(β)-bisabolene, 9.8% [4,4,13,13-2H4]-(E)-β-farnesene, and 4.2% unknowns. Competitive studies show that (3R)-nerolidyl diphosphate is the active enantiomer of (3RS)-nerolidyl diphosphate that cyclized to CDN. The kcat/K m values demonstrate that the synthase uses (E,E)-FDP as effectively as (3R)-nerolidyl diphosphate in the formation of CDN. Cyclization studies with (3R)-nerolidyl diphosphate show that the formation of CDN, (E)-β-farnesene, and β-bisabolene are enzyme dependent, but the formation of α-bisabolol in the reaction mixtures was a Mg2+-dependent solvolysis of nerolidyl diphosphate. Enzyme mechanisms are proposed for the formation of CDN from (E,E)-FDP and for the formation of CDN, (E)-β-farnesene, and β-bisabolene from (3RS)-nerolidyl diphosphate. The primary structures of cotton CDN synthase and tobacco epi-aristolochene synthase show 48% identity, suggesting similar three-dimensional structures. We used the SWISS-MODEL to test this. The two enzymes have the same overall structure consisting of two α-helical domains and epi-aristolochene synthase is a good model for the structure of CDN synthase. Several amino acids in the primary structures of both synthases superimpose. The amino acids having catalytic roles in epi-aristochene synthase are substituted in the CDN synthase and may be related to differences in catalytic properties.
Crystallization and preliminary X-ray diffraction analysis of cytochrome P450sca-2 from Streptomyces carbophilus involved in production of pravastatin sodium, a tissue-selective inhibitor of HMG-CoA reductase
Authors: Ito Shuichiro , Matsuoka Tatsuji , Watanabe Ichiro , Kagasaki Takeshi , Serizawa Nobufusa , Hata Tadashi 
Published: July 2002

Cytochrome P450sca-2 from Streptomyces carbophilus catalyzes the hydroxylation of ML-236B sodium salt to pravastatin sodium, a tissue-selective inhibitor of 3-hydroxy-3-methylglutaryl-coenzymeA (HMG-CoA) reductase. HMG-CoA reductase is a key enzyme in cholesterol biosynthesis. Crystals of the protein were obtained by the vapour-diffusion method, using ammonium sulfate as a precipitant. The crystals belong to the trigonal space group P3121 (or its enantiomorph, P3221) with unit-cell dimensions a = 103.5, c = 79.8 Å. Assuming the presence of one molecule in the asymmetric unit, the calculated value of Vm
is 2.68 Å3 Da−1. A native data set was collected to a resolution of 2.2 Å.
Crystallization and preliminary X-ray diffraction studies of undecaprenyl diphosphate synthase from Micrococcus luteus B-P 26
Authors: Fujihashi Masahiro , Shimizu Naoto , Zhang Yuan-Wei , Koyama Tanetoshi , Miki Kunio 
Published: July 2002

Undecaprenyl diphosphate synthase from Micrococcus luteus B-P 26, one of the Z-prenyl chain-elongating enzymes, was crystallized using the sitting-drop vapour-diffusion method with ammonium sulfate and lithium sulfate as precipitants. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 127.2, b = 60.2, c = 75.7 Å, β = 105.6°. The crystals diffract X-rays to at least 2.2 Å resolution using synchrotron radiation and are suitable for high-resolution crystal structure analysis.
Unfolding mechanics of holo‐ and apocalmodulin studied by the atomic force microscope
Authors: Hertadi Rukman , Ikai Atsushi 
Published: July 2002

AbstractThe structural stability of calmodulin (CaM) has been investigated previously by chemical and thermal methods. The calcium‐loaded form of CaM has been found to be exceptionally stable, because it can be exposed to temperatures >90°C or to a 9 M urea solution without a marked change in its tertiary structure, and is therefore not experimentally accessible for unfolding studies using conventional analytical methods. In this study, we have developed a system for measuring the force for mechanically unfolding CaM using an atomic force microscope (AFM) by stretching the protein from its N‐ and C‐terminal residues; we have been successful in obtaining force versus extension (F–E) curves for both apo and holo forms of CaM. In our experiment, distinguishable F–E curves have been obtained upon stretching of apoCaM and holoCaM to their full extensions. A very low force observed upon stretching of apoCaM indicated a relatively high flexibility of the apo form. On the contrary, a relatively high unfolding force and the appearance of a characteristic force peak were noted during full stretching of holoCaM. The F–E curve of the latter form of CaM most likely reflects a more rigid and probably more organized conformation of holoCaM than that of apoCaM. These experiments confirmed that the AFM is able to clearly distinguish two functionally distinct forms of CaM in terms of their mechanical properties.
Structural changes and molecular interactions of hydrophobin SC3 in solution and on a hydrophobic surface
Authors: Wang X. , Vocht M.L.De , Jonge J.De , Poolman B. , Robillard G.T. 
Published: July 2002

AbstractThe hydrophobin SC3 belongs to a class of small proteins functioning in the growth and development of fungi. Its unique amphipathic property and remarkable surface activity make it interesting not only for biological studies but also for medical and industrial applications. Biophysical studies have revealed that SC3 possesses at least three distinct conformations, named “soluble‐state SC3” for the protein in solution, and “α‐helical‐state SC3” and “β‐sheet‐state SC3” for the different states of the protein associated at a hydrophobic‐water interface. The present fluorescence study shows that the microenvironment of the dansyl‐labeled N terminus of soluble‐state SC3 is relatively hydrophobic, whereas it is hydrophilic for α‐helical‐state and β‐sheet‐state SC3. Fluorescence collisional quenching indicates that the N terminus of soluble‐state SC3 is more solvent‐accessible than those of α‐helical‐state and β‐sheet‐state SC3, with Stern‐Volmer constants for acrylamide of 4.63, 0.02, and 0.2 M−1 for the different states, respectively. Fluorescence resonance energy transfer measurements show that soluble‐state SC3 tends to associate in solution but dissociates in TFA. Fluorescence energy transfer was eliminated by conversion of soluble‐state SC3 to α‐helical‐state SC3 on a hydrophobic surface, indicating a spatial separation of the molecules in this state. By inducing the β‐sheet state, structural changes were observed, both by CD and by fluorescence, that could be fit to two exponentials with lifetimes of about 10 min and 4 h. Molecules in the β‐sheet state also underwent a slow change in spatial proximity on the hydrophobic surface, as revealed by the reappearance of fluorescence resonance energy transfer in time.
Cavities of α1‐antitrypsin that play structural and functional roles
Authors: Lee Cheolju , Maeng Jin‐Soo , Kocher Jean‐Pierre , Lee Byungkook , Yu Myeong‐Hee 
Published: July 2002

AbstractThe native form of inhibitory serine protease inhibitors (serpins) is strained, which is critical for their inhibitory activity. Previous studies on stabilizing mutations of α1‐antitrypsin, a prototype of serpins, indicated that cavities provide a structural basis for the native strain of the molecule. We have systematically mapped the cavities of α1‐antitrypsin that play such structural and functional roles by designing cavity‐filling mutations at residues that line the walls of the cavities. Results show that energetically unfavorable cavities are distributed throughout the α1‐antitrypsin molecule, and the cavity‐filling mutations stabilized the native conformation at 8 out of 10 target sites. The stabilization effect of the individual cavity‐filling mutations of α1‐antitrypsin varied (0.2–1.9 kcal/mol for each additional methylene group) and appeared to depend largely on the structural flexibility of the cavity environment. Cavity‐filling mutations that decreased inhibitory activity of α1‐antitrypsin were localized in the loop regions that interact with β‐sheet A distal from the reactive center loop. The results are consistent with the notion that β‐sheet A and the structure around it mobilize when α1‐antitrypsin forms a complex with a target protease.
The Current Status of the VOD/LD–VAC Process
Authors: Fleischer Hans J. 
Published: July 2002

Presented by C. W. Finkl
A. Finkl and Sons Company, subsidiary Republic Steel Company
The vacuum decarburization of high chromium heats called VOD in North America and LD–VAC in the rest of the world has revolutionized the melting practice of stainless steel. Since production trials started at the end of the last decade, a world production capacity of 1 720 000 tons annually has been built or is under construction. This paper deals with the latest design of the Vacuum Decarburizing Unit, and of the basic-lined slide gate ladles. It sequentially describes operating procedures giving time, chemistry, and temperatures related to the separate steps of the process. Multicombinations of melting and vacuum decarburizing facilities are explained along with cost and furnace time savings. Equipment descriptions cover cost and type of ladle refractories, vacuum pumping curve, inherent fume control, and end-point determination. Latest metallurgical developments and short cycle desulfurization in direct combination with decarburization outside the premelting furnace are also discussed, as is the advantage of vacuum ladle degassing in the VOD/LD–VAC unit.