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1.
A novel C
2-symmetric ring-fluorinated hemin, 13,17-bis(2-carboxyethyl)-2,8,12,18-tetramethyl-3,7-difluoroporphyrinatoiron(III), has
been synthesized and was incorporated into sperm whale apomyoglobin to investigate protein-induced rhombic perturbations on
the electronic structure of the active site of myoglobin (Mb) using 19F NMR spectroscopy. NMR signals for 19F atoms introduced as substituents on the present heme in ferrous low-spin and high-spin and ferric low-spin complexes have
been observed and their shifts sharply reflect not only the electronic nature of the heme iron, but also in-plane asymmetry
of the heme electronic structure. The two-fold symmetric electronic structure of the ring-fluorinated hemin is clearly manifested
in the 19F and 1H NMR spectra of its dicyano complex. The chemical equivalence of the two fluorine atoms of the heme is removed in the active
site of myoglobin and the splitting of the two 19F NMR signals provides a quantitative probe for characterizing the rhombic perturbation of the heme electronic structure induced
by the heme-protein interaction. The in-plane asymmetry of heme electronic structures in carbonmonoxy and deoxy Mbs have been
analyzed for the first time on the basis of the shift difference between the two 19F NMR signals of the heme and is interpreted in terms of iron-ligand binding and/or the orbital ground state of the heme.
A potential utility of 19F NMR, combined with the use of a symmetric fluorinated hemin, in characterizing the heme electronic structure of myoglobin
in a variety of iron oxidation, spin, and ligation states, is presented.
Received: 23 December 1999 / Accepted: 3 April 2000 相似文献
2.
3.
E. Venter A.-M. Botha 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2000,100(6):965-970
Through random amplified polymorphic DNA (RAPD) analysis we identified a putative marker linked to the Dn5 resistance gene. This marker was converted to a more reliable sequence-characterised-amplified regions (SCAR) marker. The
initial SCAR marker amplified the correct amplification product but failed to discern between the susceptible and resistant
individuals. Hence, it was utilised to sequence the internal fragment. All nested primers designed from the internal sequences
were also unable to produce any polymorphism between the susceptible and resistant cultivars. Restriction digests were then
performed on these fragments, and the restriction enzyme EcoRI was able to discern between the susceptible and resistant F2 individuals of the Dn5 population. This granted one marker amplified with the internal SCAR primer set OPF141083 the ability to differentiate between parental individuals carrying the Dn5 genes. This marker was tested in a segregating F2 population carrying the Dn5 resistance gene and proved able to differentiate between the segregating individuals. This marker may prove useful in marker
assisted selection (MAS), although performing restriction digests may hamper the throughput of a high number of samples.
Received: 4 August 1999 / Accepted: 27 August 1999 相似文献
4.
The immunoglobulin superfamily (IgSF) is an extensively diversified multigene family whose members share a common structural
feature, the Ig fold. Members of the Ig/T-cell antigen receptor (TCR) subset of the IgSF mediate antigen-specific recognition
in adaptive immune responses. Antigen-binding receptors belonging to this subset are present in all species of jawed vertebrates.
To explore whether there are additional structurally related but otherwise distinct members of this subset, we have developed
a technique termed the short-primer polymerase chain reaction (PCR) that targets structurally conserved short motifs in the
Ig fold. Large-scale sequencing efforts and recent advances in information biotechnology, including "electronic PCR," provide
additional computational means to implement similarly directed searches within databases. The use of these approaches has
led to the discoveries of Ig/TCR homologues in a variety of phylogenetically diverse organisms, a diversified family of novel
immune-type receptor genes, as well as a novel human IgSF member. The potential of random sequencing efforts and virtual screening
of databases is described in the context of two novel genes in bony fish. The various methodologies that are discussed and
the examples shown provide means for further investigating, and/or elucidating novel, IgSF receptors as well as components
of pathways that are involved in immune responses in both traditional and nontraditional model systems. 相似文献
5.
Binding of divalent magnesium to RNA homopolymers was evaluated by titration calorimetry and analyzed in terms of the McGhee
and von Hippel model of a one-dimensional infinite lattice. Examination of Mg2+ binding data for ds poly(A)×poly(U), ss poly(A), and ss poly(U) revealed that the sign of the enthalpy term relates to the
mode of metal binding (inner or outer sphere), which in turn is directly related to the structural ordering of the polynucleotide
in solution. This approach provides details of the thermodynamics of metal binding, stoichiometry, and coordination chemistry
that underlie the structural and catalytic chemistry of RNA and offers a potential new probe of the solution structure of
nucleic acids. 相似文献
6.
7.
Summary. Ten years after the establishment of the term proteome, the science surrounding it has yet to fulfill its potential. While
a host of technologies have generated lists of protein names, there are only a few reported studies that have examined the
individual proteins at the covalent chemical level defined as protein species in 1997 and their function. In the current study,
we demonstrate that this is possible with two-dimensional gel electrophoresis (2-DE) and mass spectrometry by presenting clear
evidence of in vivo N-terminal alpha A crystallin truncation and relating this newly detected protein species to alpha crystallin
activity regulation by protease cleavage in the healthy young murine lens. We assess the present state of technology and suggest
a shift in resources and paradigm for the routine attainment of the protein species level in proteomics. 相似文献
8.
The negative gravitropism of the sporangiophores of Phycomyces blakesleeanus Burgeff is elicited by different sensory inputs, which include flexure of the growing zone, buoyance of lipid globules and
sedimentation of paracrystalline proteins, so-called octahedral crystals (C. Schimek et al., 1999a, Planta 210: 132–142).
Gravity-induced absorbance changes (GIACs), which are associated with primary events of gravity sensing, were detected in
the growing zones of sporangiophores. After placing sporangiophores horizontally, GIACs were detected after a latency of about
5 min, i.e. 15–25 min prior to gravitropic bending. The spectroscopic properties of the GIACs indicate that gravitropic stimulation
could imply the reduction of cytochromes. The GIACs were spectrally distinct from light-induced absorbance changes (LIACs),
showing that the primary responses of the light and gravity transduction chains are different. A dual stimulation with gravity
and light generated GIAC-LIACs which were distinct from the absorbance changes occurring after the single stimuli and which
indicate that light and gravity interact early in the respective transduction chains.
Received: 2 September 1999 / Accepted: 9 November 1999 相似文献
9.
R. E. Voorrips M. C. Jongerius H. J. Kanne 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1997,94(1):75-82
A genetic map covering 615 cM in 12 linkage groups was assembled based on 92 RFLP and AFLP markers segregating in a population
of 107 doubled haploid lines (DH lines) of Brassica oleracea. The DH-line population was obtained through microspore culture from the of two homozygous parents: DH-line Bi derived from the cabbage landrace Bindsachsener, and DH-line Gr from broccoli cv ‘Greenia’.
Sixty-five percent of the loci, and in some cases complete linkage groups, displayed distorted segregation ratios, a frequency
much higher than that observed in populations of the same species. DH-line Bi was resistant to clubroot, which is caused by a Dutch field isolate of Plasmodiophora brassicae. Resistance in the DH-line population was determined in two ways: by assigning symptom grades to each plant, and by measuring
the fresh weights of the healthy and affected parts of the root system of each plant. Using a multiple QTL mapping approach
to analyze the fresh weight data, we found two loci for clubroot resistance; these were designated pb-3 and pb-4. The additive effects of these loci were responsible for 68% of the difference between the parents and for 60% of the genetic
variance among DH-line means. Also, indications for the presence of two additional, minor QTLs were found. Analysis of symptom
grades revealed the two QTLs pb-3 and pb-4, as well as one of the two minor QTLs indicated by analysis of the fresh weight data.
Received: 29 April 1996 / Accepted: 10 May 1996 相似文献
10.
Summary Application of 1H 2D NMR methods to solubilized membrane proteins and peptides has up to now required the use of selectively deuterated detergents. The unavailability of any of the common biochemical detergents in deuterated form has therefore limited to some extent the scope of this approach. Here a 1H NMR method is described which allows structure determination of membrane peptides and small membrane proteins by 1H 2D NMR in any type of non-deuterated detergent. The approach is based on regioselective excitation of protein resonances with DANTE-Z or spin-pinging pulse trains. It is shown that regioselective excitation of the amide-aromatic region of solubilized membrane proteins and peptides leads to an almost complete suppression of the two orders of magnitude higher contribution of the protonated detergent to the 1H NMR spectrum. Consistently TOCSY, COSY and NOESY sequences incorporating such regioselective excitation in the F2 dimension yield protein 1H 2D NMR spectra of quality comparable to those obtained in deuterated detergents. Regioselective TOCSY and NOESY spectra display all through-bond and through-space correlations within amide-aromatic protons and between these protons and aliphatic and -protons. Regioselective COSY spectra provide scalar coupling constants between amide and -protons. Application of the method to the membrane-active peptide mastoparan X, solubilized in n-octylglucoside, yields complete sequence-specific assignments and extensive secondary structure-related spatial proximities and coupling constants. It is shown that mastoparan adopts an -helical conformation when bound to nonionic detergent micelles. The present method is expected to increase the applicability of 1H solution NMR methods to membrane proteins and peptides.Abbreviations 2D NMR
two-dimensional NMR
- COSY
correlated spectroscopy
- DANTE
delays alternating nutations for tailored excitation
- NOESY
nuclear Overhauser enhancement spectroscopy
- TOCSY
total correlation spectroscopy 相似文献
11.
Krabben L van Rossum BJ Castellani F Bocharov E Schulga AA Arseniev AS Weise C Hucho F Oschkinat H 《FEBS letters》2004,564(3):319-324
Solid-state magic-angle spinning nuclear magnetic resonance (NMR) has sufficient resolving power for full assignment of resonances and structure determination of immobilised biological samples as was recently shown for a small microcrystalline protein. In this work, we show that highly resolved spectra may be obtained from a system composed of a receptor-toxin complex. The NMR sample used for our studies consists of a membrane preparation of the nicotinic acetylcholine receptor from the electric organ of Torpedo californica which was incubated with uniformly 13C-,15N-labelled neurotoxin II. Despite the large size of the ligand-receptor complex ( > 290 kDa) and the high lipid content of the sample, we were able to detect and identify residues from the ligand. The comparison with solution NMR data of the free toxin indicates that its overall structure is very similar when bound to the receptor, but significant changes were observed for one isoleucine. 相似文献
12.
Although the significance of canopy plant communities to ecosystem function is well documented, the process by which such
communities become established in trees remains poorly known. Colonization of tree surfaces by canopy-dwelling plants often
begins with the establishment of bryophytes, so the conditions that affect the dispersal of bryophytes in the forest canopy
merit study. We assessed success rates of one mechanism of bryophyte propagation, the aerial dispersal of macroscopic fragments,
using an experimental approach. We quantified interception and retention of marked fragments released from a 36 cm×36 cm grid
50 cm above branches of saplings and mature trees of the species Ocotea tonduzii in a montane cloud forest in Costa Rica. Only 1% of bryophyte fragments dropped over sapling crowns in this manner were retained
for the 6-month duration of the study, while branches in the forest canopy with intact epiphyte loads and branches that had
been stripped of their epiphytes retained 24% and 5%, respectively. Our results suggest that larger-diameter branches and
the presence of other epiphytes can both improve the retention of bryophyte fragments on canopy branches. Further work will
be needed to address the relative roles of other dispersal mechanisms (spores, gemmae, microscopic bryophyte fragments) and
the dynamics of growth and establishment of macroscopic bryophyte fragments following their interception.
Received: 1 March 1999 / Accepted: 24 August 1999 相似文献
13.
14.
Daniel Haydon Susan Lea Liz Fry Nick Knowles Alan R. Samuel David Stuart Mark E. J. Woolhouse 《Journal of molecular evolution》1998,46(4):465-475
The VP1 capsid protein of foot and mouth disease virus (FMDV) is highly polymorphic and contains several of the major immunogenic
sites important to effective antibody neutralization and subsequent viral clearance by the immune system. Whether this high
level of polymorphism is of adaptive value to the virus remains unknown. In this study we examined sequence data from a set
of 55 isolates in order to establish the nature of selective pressures acting on this gene. Using the known molecular structure
of VP1, the rates and ratios of different types of nonsynonymous and synonymous changes were compared between different parts
of the protein. All parts of the protein are subject to purifying selection, but this is greatest amongst those amino acid
residues within β-strands and is significantly reduced at residues exposed on the capsid surface, which include those residues
demonstrated by previous mutational analyses to permit the virus to escape from monoclonal antibody binding. The ratios of
nonsynonymous substitution resulting in various forms of physicochemically radical and conserved amino acid change were shown
to be largely equal throughout these different parts of the protein. There was a consistently higher level of nonsynonymous
and charge radical sites in those regions of the gene coding for residues exposed on the outer surface of the capsid and a
marked difference in the use of amino acids between surface and nonsurface regions of the protein. However, the analysis is
consistent with the hypothesis that the observed sequence variation arises where it is least likely to be disruptive to the
higher-order structure of the protein and is not necessarily due to positive Darwinian selection.
Received: 8 March 1997 / Accepted: 12 August 1997 相似文献
15.
B. Riegl 《Coral reefs (Online)》1999,18(1):63-73
Reef coral communities in a non-reef setting on shallow, flat hardgrounds were quantitatively sampled in Dubai Emirate (UAE,
Southern Arabian Gulf) before and after a coral mass mortality in 1996. The coral fauna consisted of 34 scleractinian species
before and 27 after the event, which removed virtually all Acropora. No alcyonacea were recorded. Five community types were identified and characterized by the dominant species: (A) a sparse
Porites lutea community in sandy areas, (B) a dense Acropora clathrata community in areas with little sand, (C) a faviid community in muddy areas, (D) a Siderastrea savignyana community in sandy areas, and (E) a Porites compressa community, which built a framework in sandy areas. These communities are comparable to those described from other areas of
the Gulf, where a stable pattern of community differentiation appears to exist. The spatial distribution and dynamics of the
coral communities appears to be strongly influenced by mass mortality events recurring every 15 to 20 y. A combination of
extreme water temperatures and high sedimentation/turbidity appear to be the major cause of mortality.
Accepted: 13 June 1998 相似文献
16.
Peter J. Simmons F. Claire Rind 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》1997,180(3):203-214
The LGMD2 belongs to a group of giant movement-detecting neurones which have fan-shaped arbors in the lobula of the locust
optic lobe and respond to movements of objects. One of these neurones, the LGMD1, has been shown to respond directionally
to movements of objects in depth, generating vigorous, maintained spike discharges during object approach. Here we compare
the responses of the LGMD2 neurone with those of the LGMD1 to simulated movements of objects in depth and examine different
image cues which could allow the LGMD2 to distinguish approaching from receding objects. In the absence of stimulation, the
LGMD2 has a resting discharge of 10–40 spikes s−1 compared with <1 spike s−1 for the LGMD1. The most powerful excitatory stimulus for the LGMD2 is a dark object approaching the eye. Responses to approaching
objects are suppressed by wide field movements of the background. Unlike the LGMD1, the LGMD2 is not excited by the approach
of light objects; it specifically responds to movement of edges in the light to dark direction. Both neurones rely on the
same monocular image cues to distinguish approaching from receding objects: an increase in the velocity with which edges of
images travel over the eye; and an increase in the extent of edges in the image during approach.
Accepted: 23 October 1996 相似文献
17.
B. J. Goodfellow A. L. Macedo Pedro Rodrigues Isabel Moura Victor Wray José J. G. Moura 《Journal of biological inorganic chemistry》1999,4(4):421-430
The use of standard 2D NMR experiments in combination with 1D NOE experiments allowed the assignment of 51 of the 58 spin systems of oxidised [3Fe-4S] ferredoxin isolated from Desulfovibrio gigas. The NMR solution structure was determined using data from 1D NOE and 2D NOESY spectra, as distance constraints, and information from the X-ray structure for the spin systems not detected by NMR in torsion angle dynamics calculations to produce a family of 15 low target function structures. The quality of the NMR family, as judged by the backbone r.m.s.d. values, was good (0.80?Å), with the majority of φ/ψ angles falling within the allowed region of the Ramachandran plot. A comparison with the X-ray structure indicated that the overall global fold is very similar in solution and in the solid state. The determination of the solution structure of ferredoxin II (FdII) in the oxidised state (FdIIox) opens the way for the determination of the solution structure of the redox intermediate state of FdII (FdIIint), for which no X-ray structure is available. 相似文献
18.
Plant species diversity as a driver of early succession in abandoned fields: a multi-site approach 总被引:15,自引:0,他引:15
W. H. Van der Putten S. R. Mortimer K. Hedlund C. Van Dijk V. K. Brown J. Lepä C. Rodriguez-Barrueco J. Roy T. A. Diaz Len D. Gormsen G. W. Korthals S. Lavorel I. Santa Regina P. Smilauer 《Oecologia》2000,124(1):91-99
Succession is one of the most studied processes in ecology and succession theory provides strong predictability. However,
few attempts have been made to influence the course of succession thereby testing the hypothesis that passing through one
stage is essential before entering the next one. At each stage of succession ecosystem processes may be affected by the diversity
of species present, but there is little empirical evidence showing that plant species diversity may affect succession. On
ex-arable land, a major constraint of vegetation succession is the dominance of perennial early-successional (arable weed)
species. Our aim was to change the initial vegetation succession by the direct sowing of later-successional plant species.
The hypothesis was tested that a diverse plant species mixture would be more successful in weed suppression than species-poor
mixtures. In order to provide a robust test including a wide range of environmental conditions and plant species, experiments
were carried out at five sites across Europe. At each site, an identical experiment was set up, albeit that the plant species
composition of the sown mixtures differed from site to site. Results of the 2-year study showed that diverse plant species
mixtures were more effective at reducing the number of natural colonisers (mainly weeds from the seed bank) than the average
low-diversity treatment. However, the effect of the low-diversity treatment depended on the composition of the species mixture.
Thus, the effect of enhanced species diversity strongly depended on the species composition of the low-diversity treatments
used for comparison. The effects of high-diversity plant species mixtures on weed suppression differed between sites. Low-productivity
sites gave the weakest response to the diversity treatments. These differences among sites did not change the general pattern.
The present results have implications for understanding biological invasions. It has been hypothesised that alien species
are more likely to invade species-poor communities than communities with high diversity. However, our results show that the
identity of the local species matters. This may explain, at least partly, controversial results of studies on the relation
between local diversity and the probability of being invaded by aliens.
Received: 13 July 1999 / Accepted: 4 February 2000 相似文献
19.
Chee Choy Kok Emma J. Croager Campbell S. Witt Lynette Kiers Frank L. Mastaglia Lawrence J. Abraham M. J. Garlepp 《Immunogenetics》1999,49(6):508-516
Inclusion body myositis (IBM) is a form of idiopathic inflammatory myopathy of unknown aetiology. A strong association with
HLA class II (HLA-DR3) suggested a role for genes in the human major histocompatibility complex (MHC) in the predisposition to this disease. In
this study, we have taken advantage of the ancestral haplotype (AH) concept and historical recombinations to map for a possible
susceptibility gene(s) in the MHC. We performed detailed typing of three MHC-related HSP70 genes and defined allelic combinations in the context of MHC AH. We also modified existing methods to give a simple and accurate
method for typing two TNF microsatellites. Using the HSP70 and TNF markers and HLA-DR, –B, and C4 typing of our patients with IBM, we defined a potential site for the MHC-associated susceptibility gene(s) in the region
between HLA-DR and C4.
Received: 16 July 1998 / Revised: 14 January 1999 相似文献
20.
Shigetoshi Aono I. Bertini J. A. Cowan Claudio Luchinat Antonio Rosato Maria Silvia Viezzoli 《Journal of biological inorganic chemistry》1996,1(6):523-528
The oxidized Fe7S8 ferredoxin from Bacillus schlegelii, containing both [Fe3S4]+ and [Fe4S4]2+ clusters, has been investigated by 1H NMR spectroscopy. An extensive sequence-specific assignment of the hyperfine-shifted resonances has been obtained by making
use of a computer-generated structural model. The pattern and the temperature dependence of the hyperfine shifts of the β-CH2 protons of the cysteines coordinating the [Fe3S4]+ cluster are rationalized in terms of magnetic interactions between the iron ions. The same approach holds for the hyperfine
coupling with 57Fe. It is shown that the magnetic interactions are more asymmetric in Fe7S8 ferredoxins than in Fe3S4 ferredoxins. The NMR non-observability of the β-CH2 protons of coordinated cysteines in the one-electron-reduced form has been discussed.
Received: 19 June 1996 / Accepted: 2 August 1996 相似文献