scholarly journals Phenylalanine ammonia-lyase (PAL) from tobacco (Nicotiana tabacum): characterization of the four tobacco PAL genes and active heterotetrameric enzymes1

2009 ◽  
Vol 424 (2) ◽  
pp. 233-242 ◽  
Author(s):  
Angelika I. Reichert ◽  
Xian-Zhi He ◽  
Richard A. Dixon
Keyword(s):  
Nicotiana Tabacum ◽  
Phenylalanine Ammonia Lyase ◽  
Consensus Sequence ◽  
Three Dimensional ◽  
Pcr Amplification ◽  
Dimensional Structure ◽  
Kinetic Properties ◽  
Cdna Clones ◽  
Phenylpropanoid Biosynthesis ◽  
The Individual

PAL (L-phenylalanine ammonia-lyase), the first enzyme of phenylpropanoid biosynthesis, is often encoded by multigene families in plants. A PCR-based approach was used to isolate cDNA clones corresponding to the four PAL genes of tobacco (Nicotiana tabacum). By careful comparison of cDNA and genomic clones, a new PAL gene (PAL4) was defined. PCR amplification of PAL sequences from cDNA led to the generation of chimaeric clones between PAL1 and PAL4, and incorrect annotation of PAL4 ESTs (expressed sequence tags) as PAL1 in the EST database has given rise to a randomly shuffled tentative consensus sequence. The PAL2 previously described in the literature was shown, by domain swapping experiments with PAL1, to possess a single nucleotide substitution leading to an inactive enzyme. The altered amino acid resulting from this substitution maps to the base of the active site pocket in the three-dimensional structure of PAL. The inactive PAL2 allele could not be recovered from 13 different tobacco cultivars examined. PALs 1–4 were co-expressed in multiple plant organs, and were also co-induced following exposure of cell cultures to yeast elicitor or methyl jasmonate. All four tobacco PAL proteins expressed in Escherichia coli displayed normal Michaelis–Menten kinetics, with Km values between 36 and 60 μM. Co-expression of different PAL proteins in E. coli resulted in formation of heterotetramers, which possessed kinetic properties within the same range as those of the individual homotetramers. The potential physiological function of heterotetrameric PAL forms is discussed.

scholarly journals Crystal structure of 2-methyl-4-[(thiophen-2-yl)methylidene]-1,3-oxazol-5(4H)-one

2015 ◽  
Vol 71 (2) ◽  
pp. o123-o124 ◽  
Author(s):  
Preetika Sharma ◽  
K. N. Subbulakshmi ◽  
B. Narayana ◽  
K. Byrappa ◽  
Rajni Kant
Keyword(s):  
Crystal Structure ◽  
Hydrogen Bonds ◽  
Title Compound ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Asymmetric Unit ◽  
Π Interactions ◽  
Compound C ◽  
Independent Molecules ◽  
The Individual

The asymmetric unit of the title compound, C9H7NO2S, contains two crystallographically independent molecules (AandB). Both molecules are almost planar [maximum deviations = 0.047 (1) and 0.090 (1) Å, respectively, for the S atoms] with the oxazole and thiophene rings being inclined to one another by 2.65 (16)° in moleculeAand by 4.55 (15)° in moleculeB. In the crystal, the individual molecules are linkedviaC—H...O hydrogen bonds, forming –A–B–A–B– chains along the [10-1] direction. The chains are linkedviaC—H...π and π–π interactions [intercentroid distances = 3.767 (2) and 3.867 (2) Å] involving inversion-related oxazole and thiophene rings in both molecules, forming a three-dimensional structure.

scholarly journals The 3D morphology of the ejecta surrounding VY Canis Majoris

2007 ◽  
Vol 3 (S242) ◽  
pp. 256-260
Author(s):  
Terry Jay Jones ◽  
Roberta M. Humphreys ◽  
L. Andrew Helton
Keyword(s):  
Three Dimensional ◽  
Independent Method ◽  
Dimensional Structure ◽  
Line Of Sight ◽  
Circumstellar Dust ◽  
Geometric Objects ◽  
Imaging Polarimetry ◽  
Intensity Image ◽  
The Individual ◽  
Dust Distribution

AbstractWe use second epoch images taken with WFPC2 on the HST and imaging polarimetry taken with the HST/ACS/HRC to explore the three dimensional structure of the circumstellar dust distribution around the red supergiant VY Canis Majoris. Transverse motions, combined with radial velocities, provide a picture of the kinematics of the ejecta, including the total space motions. The fractional polarization and photometric colors provide an independent method of locating the physical position of the dust along the line-of-sight. Most of the individual arc-like features and clumps seen in the intensity image are also features in the fractional polarization map, and must be distinct geometric objects. The location of these features in the ejecta of VY CMa using kinematics and polarimetry agree well with each other, and strongly suggest they are the result of relatively massive ejections, probably associated with magnetic fields.

scholarly journals Interactions and three-dimensional localization of a group of nuclear pore complex proteins.

1994 ◽  
Vol 126 (3) ◽  
pp. 603-617 ◽  
Author(s):  
N Panté ◽  
R Bastos ◽  
I McMorrow ◽  
B Burke ◽  
U Aebi
Keyword(s):  
Nuclear Pore Complex ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Nuclear Pore ◽  
Western Blots ◽  
Cytoplasmic Filaments ◽  
Quick Freezing ◽  
The Individual ◽  
Antibody Labeling

We have used antibodies directed against a number of nuclear pore complex (NPC) proteins to determine their mutual interactions and location within the three-dimensional structure of the NPC. A monoclonal antibody, termed QE5, recognized three NPC polypeptides, p250, NUP153, and p62 on Western blots, and labeled the nuclear envelope of several cultured cell lines by immunofluorescence microscopy. These three polypeptides contained O-linked N-acetylglucosamine residues and were released from the NPC by detergent/high-salt treatment as discrete high molecular weight complexes. p250 was found in association with a novel 75 kD protein, NUP153 was released as a homo-oligomer of about 1 megadalton, and p62 was associated with polypeptides of 58 and 54 kD (previously reported by Finlay, D. R., E. Meier, P. Bradley, J. Horecka, and D. J. Forbes. 1991. J. Cell Biol. 114:169-183). p75, p58, and p54 were not galactosylated in vitro. Xenopus oocyte NEs were labeled with gold-conjugated QE5 and prepared for electron microscopy by quick freezing/freeze drying/rotary metal shadowing. This EM preparation method enabled us to more precisely localize the epitopes of this antibody to the cytoplasmic filaments and the nuclear basket of the NPC. Since QE5 recognizes three O-linked NPC glycoproteins, its labeling was compared with that of the lectin wheat germ agglutinin which recognizes O-linked N-acetylglucosamine moieties. The two probes were found to yield similar, although not identical, distributions of label. To identify the individual proteins with particular NPC components, we have used an anti-peptide antibody against NUP153 and a monospecific anti-p250 polyclonal antibody. Labeling with these two antibodies has documented that NUP153 is a constituent of the nuclear basket with at least one of its epitopes residing in its terminal ring, whereas p250 is a constituent of the cytoplasmic filaments.

scholarly journals An open reading frame in intron seven of the sea urchin DNA-methyltransferase gene codes for a functional AP1 endonuclease

2002 ◽  
Vol 365 (3) ◽  
pp. 833-840 ◽  
Author(s):  
Anna Valentina CIOFFI ◽  
Diana FERRARA ◽  
Maria Vittoria CUBELLIS ◽  
Francesco ANIELLO ◽  
Marcella CORRADO ◽  
...  
Keyword(s):  
Sea Urchin ◽  
Dna Methyltransferase ◽  
Developmental Stages ◽  
Genome Structure ◽  
Three Dimensional ◽  
Paracentrotus Lividus ◽  
Open Reading Frame ◽  
Dimensional Structure ◽  
Cdna Clones ◽  
Reading Frame

Analysis of the genome structure of the Paracentrotus lividus (sea urchin) DNA methyltransferase (DNA MTase) gene showed the presence of an open reading frame, named METEX, in intron 7 of the gene. METEX expression is developmentally regulated, showing no correlation with DNA MTase expression. In fact, DNA MTase transcripts are present at high concentrations in the early developmental stages, while METEX is expressed at late stages of development. Two METEX cDNA clones (Met1 and Met2) that are different in the 3′ end have been isolated in a cDNA library screening. The putative translated protein from Met2 cDNA clone showed similarity with Escherichia coli endonuclease III on the basis of sequence and predictive three-dimensional structure. The protein, overexpressed in E. coli and purified, had functional properties similar to the endonuclease specific for apurinic/apyrimidinic (AP) sites on the basis of the lyase activity. Therefore the open reading frame, present in intron 7 of the P. lividus DNA MTase gene, codes for a functional AP endonuclease designated SuAP1.

Comparison of IVEM and quick-freeze, deep-etch, rotary shadow images of the cell cytoplasm

1993 ◽  
Vol 51 ◽  
pp. 66-67
Author(s):  
Carolyn A. Larabell
Keyword(s):  
Electron Microscopy ◽  
Three Dimensional ◽  
Freeze Fracture ◽  
Dimensional Structure ◽  
Whole Mount ◽  
Thin Section Electron Microscopy ◽  
Section Electron ◽  
Excellent Method ◽  
Fine Structures ◽  
The Individual

The techniques of freeze-fracture and quick-freeze, deep-etch, rotary-shadow electron microscopy have provided information about cellular structures that cannot be obtained from thin section electron microscopy. Freeze-fracture replicas provide extensive views of membranes and the arrangement of integral membrane proteins, but filamentous structures such as the cytoskeleton and extracellular matrix are not visualized because they are embedded in ice. In order to visualize such fine structures, it is necessary to deeply etch (or freeze-dry) cells that have been rapidly frozen, thus revealing about 0.1 to 0.3 μm of three dimensional structure. Replicas of cells prepared in this fashion provide excellent high magnification views of the individual filaments and their interactions with the plasma membrane as well as with membrane-bounded organelles. Another excellent method for visualization of cytoskeletal structures is whole mount electron microscopy. In this paper, I describe views of the egg cytoskeleton obtained from visualization of the isolated egg cortex prepared as a whole mount at 400 kV and compare these results with those obtained from replicas of cells that have been rapidly frozen men fractured and deeply etched.

scholarly journals The Molecular Architecture of Galactose Mutarotase/UDP-Galactose 4-Epimerase from Saccharomyces cerevisiae

2005 ◽  
Vol 280 (23) ◽  
pp. 21900-21907 ◽  
Author(s):  
James B. Thoden ◽  
Hazel M. Holden
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Active Sites ◽  
Polypeptide Chain ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Dual Function ◽  
Molecular Architecture ◽  
Single Polypeptide Chain ◽  
Leloir Pathway ◽  
The Individual

The metabolic pathway by which β-d-galactose is converted to glucose 1-phosphate is known as the Leloir pathway and consists of four enzymes. In most organisms, these enzymes appear to exist as soluble entities in the cytoplasm. In yeast such as Saccharomyces cerevisiae, however, the first and last enzymes of the pathway, galactose mutarotase and UDP-galactose 4-epimerase, are contained within a single polypeptide chain referred to as Gal10p. Here we report the three-dimensional structure of Gal10p in complex with NAD+, UDP-glucose, and β-d-galactose determined to 1.85-Å resolution. The enzyme is dimeric with dimensions of ∼91 Å × 135 Å × 108 Å and assumes an almost V-shaped appearance. The overall architecture of the individual subunits can be described in terms of two separate N- and C-terminal domains connected by a Type II turn formed by Leu-357 to Val-360. The first 356 residues of Gal10p fold into the classical bilobal topology observed for all other UDP-galactose 4-epimerases studied thus far. This N-terminal domain contains the binding sites for NAD+ and UDP-glucose. The polypeptide chain extending from Glu-361 to Ser-699 adopts a β-sandwich motif and harbors the binding site for β-d-galactose. The two active sites of Gal10p are separated by over 50 Å. This investigation represents the first structural analysis of a dual function enzyme in the Leloir pathway.

scholarly journals Functional and molecular modelling studies of two hereditary fructose intolerance-causing mutations at arginine 303 in human liver aldolase

2000 ◽  
Vol 350 (3) ◽  
pp. 823-828 ◽  
Author(s):  
Rita SANTAMARIA ◽  
Gabriella ESPOSITO ◽  
Luigi VITAGLIANO ◽  
Vincenza RACE ◽  
Immacolata PAGLIONICO ◽  
...  
Keyword(s):  
Homology Modelling ◽  
Three Dimensional ◽  
Catalytic Efficiency ◽  
Dimensional Structure ◽  
Kinetic Properties ◽  
Wild Type ◽  
Hereditary Fructose Intolerance ◽  
Fructose Intolerance ◽  
Aldolase B ◽  
Fructose 1,6 Bisphosphate

We have identified a novel hereditary fructose intolerance mutation in the aldolase B gene (i.e. liver aldolase) that causes an arginine-to-glutamine substitution at residue 303 (Arg303 → Gln). We previously described another mutation (Arg303 → Trp) at the same residue. We have expressed the wild-type protein and the two mutated proteins and characterized their kinetic properties. The catalytic efficiency of protein Gln303 is approx. 1/100 that of the wild-type for substrates fructose 1,6-bisphosphate and fructose 1-phosphate. The Trp303 enzyme has a catalytic efficiency approx. 1/4800 that of the wild-type for fructose 1,6-bisphosphate; no activity was detected with fructose 1-phosphate. The mutation Arg303 → Trp thus substitution impairs enzyme activity more than Arg303 → Gln. Three-dimensional models of wild-type, Trp303 and Gln303 aldolase B generated by homology-modelling techniques suggest that, because of its larger size, tryptophan exerts a greater deranging effect than glutamine on the enzyme's three-dimensional structure. Our results show that the Arg303 → Gln substitution is a novel mutation causing hereditary fructose intolerance and provide a functional demonstration that Arg303, a conserved residue in all vertebrate aldolases, has a dominant role in substrate binding during enzyme catalysis.

scholarly journals The interpretation of structure from motion

1979 ◽  
Vol 203 (1153) ◽  
pp. 405-426 ◽  
Keyword(s):  
Structure From Motion ◽  
Three Dimensional ◽  
Point Of View ◽  
Dimensional Structure ◽  
Two Dimensional ◽  
Three Dimensional Structure ◽  
Structure And Motion ◽  
Rigid Objects ◽  
The Individual

The interpretation of structure from motion is examined from a computional point of view. The question addressed is how the three dimen­sional structure and motion of objects can be inferred from the two dimensional transformations of their projected images when no three dimensional information is conveyed by the individual projections. The following scheme is proposed: (i) divide the image into groups of four elements each; (ii) test each group for a rigid interpretation; (iii) combine the results obtained in (ii). It is shown that this scheme will correctly decompose scenes containing arbitrary rigid objects in motion, recovering their three dimensional structure and motion. The analysis is based primarily on the ʻstructure from motion’ theorem which states that the structure of four non-coplanar points is recoverable from three orthographic projections. The interpretation scheme is extended to cover perspective projections, and its psychological relevance is discussed.

APPLICATION OF TERRESTRIAL PHOTOGRAMMETRY TO THE CREATION OF A 3D MODEL OF THE SAINT HEDWIG CHAPEL IN THE KAŇOVICE

2014 ◽  
Vol 40 (1) ◽  
pp. 8-13
Author(s):  
Milan Mikoláš ◽  
Petr Jadviščok ◽  
Vlastimil Molčák
Keyword(s):  
Camera Calibration ◽  
3D Model ◽  
Three Dimensional ◽  
Digital Camera ◽  
Dimensional Structure ◽  
Building Structures ◽  
Whole Process ◽  
The Creation ◽  
The Individual ◽  
Terrestrial Photogrammetry

The present article focuses on application of terrestrial photogrammetry for the purposes of creation of photogrammetric documentation of building structures with the use of digital camera – a widely accessible device. First, the article briefly describes the individual intermediate operations of the whole process leading to the creation of a three-dimensional structure. Next, attention is given to operations related to camera calibration, reconnaissance of the locality of interest, photographing itself, creation of the 3D model as well as to presentation of graphical output. In conclusion, the article focuses on determining the accuracy of photogrammetric measuring.

The Structure and Subunit Organization of the Bacterial Flagellar Motor by Scanning Transmission Electron Microscopy and Cryoelectron Microscopy

1990 ◽  
Vol 48 (1) ◽  
pp. 278-279
Author(s):  
Gina E. Sosinsky ◽  
Noreen R. Francis ◽  
Charles D. DeRosier ◽  
David J. DeRosier ◽  
James Hainfeld ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Basal Body ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Cryoelectron Microscopy ◽  
Data Set ◽  
Bacterial Flagellum ◽  
The Individual ◽  
The Masses ◽  
Subunit Organization

The bacterial flagellum is unique in having a rotary motor. In Salmonella typhimurium, the basal body, a component of the motor, consists of four rings (denoted M, S, L, and P) threaded on a coaxial rod. The M, L, and P rings are each composed of a different protein: FliF=61 kD, FlgH=22 kD, and FlgI=36 kD, respectively. The rod contains at least four different proteins: FlgB=15 kD, FlgC=14 kD, FlgF=26 kD, and FlgG=28 kD. Using quantitative gel analysis, Jones et al. estimated that there are about 26 copies of FlgG, FlgH, Flgl and FliF, and 6 copies of FlgB, FlgC and FlgF per basal body. The total mass of these 7 proteins per basal body is ∽4200 kD. There appear to be additional proteins in the basal body, but their locations and amounts are not known. Our aim is to produce subcomplexes of the basal body and determine their structures and masses using electron microscopy. This approach is complementary to that of Jones et al. and can reveal the presence and amounts of as yet unidentified components. We find, in pH3- or pH4-treated preparations of basal bodies, four subcomplexes of the hook basal body complex (HBB): the HLPRS (hook, L and P rings on the distal rod, proximal rod, S ring); the HLPR (lacks the M and S rings), the HLP (lacks the M, S, and proximal rod); and the LP complex (Figs. 1 and 2). We have been able to visualize the three-dimensional structure and the subunit organization using the combined techniques of cryoelectron microscopy and image analysis. These studies suggest that the S ring is a separate component from the rod or M ring and that the rod consists of two sections. Because the different sub-complexes are distinguishable in a field of particles, we measured the molecular masses of the individual subcomplexes using the Brookhaven STEM even though these preparations are not homogeneous (Fig. 3). All the structures analyzed so far had hooks attached. We measured the length and mass/length from STEM images and then subtracted the mass of the hook. Preliminary results show that the molecular mass of the hookless basal body is 4400−500 kD (n=165), that of the LP-rod (proximal and distal) is 3500±300 kD (n=52), and that of the LP-distal rod is 2300±450 kD (n=76) (Fig. 4). The difference between these three molecular weights gives estimates of the mass of the M and S rings (4400 - 3500 = 900 kD) and proximal rod, 3500 − 2300 = 1200 kD. The mass of the M and S rings may be underestimated due to the undetected presence of HLPRS subcomplexes in the basal body data set. We are presently measuring and re-evaluating masses for the subcomplexes in order to get more accurate estimates of the masses and numbers of subunits.

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