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1.
G. Musci Terri Z. L. Fraterrigo Lilia Calabrese David R. McMillin 《Journal of biological inorganic chemistry》1999,4(4):441-446
The possibility that ceruloplasmin (CP) functions as a copper transferase has fueled a continuing interest in studies of
the copper release process. The principal goal of the current investigation has been to identify the most labile copper centers
in sheep protein. In fact, subjecting the enzyme to a slow flux of cyanide at pH 5.2 under nitrogen in the presence of ascorbate
and a phenanthroline ligand produces partially demetalated forms of the protein. By standard chromatographic techniques it
is possible to isolate protein with a Cu/CP ratio of ∼4 or ∼5 as opposed to the native protein which has Cu/CP=5.8. In contrast
to other blue oxidases, analysis suggests that CP preferentially loses its type 1 coppers under these conditions. Thus, the
spectroscopic signals from the type 1 centers exhibit a loss of intensity while the EPR signal of the type 2 copper becomes
stronger. Furthermore, the Cu/CP≈4 and Cu/CP≈5 components retain about 50% of the activity of the native protein, consistent
with an intact type 2/type 3 cluster. All three type 1 copper sites appear to suffer copper loss. Reconstitution with a copper(I)
reagent restores the spectroscopic properties of the native protein and 90% of the original activity. The results suggest
a possible functional significance for the presence of three type 1 coppers in CP. By employing a pool of redox-active but
relatively labile type 1 copper centers, the enzyme can serve as a copper donor, if necessary, without completely sacrificing
its oxidase activity.
Received: 15 February 1999 / Accepted: 22 April 1999 相似文献
2.
The involvement of amino acid residues previously proposed on the basis of structural data to have roles in the ferroxidase and diamine oxidase activities of human ceruloplasmin was investigated. Variants of human ceruloplasmin, in which residues proposed to be involved in electron transfer and/or iron-binding had been altered by site-directed mutagenesis, were expressed in HEK293 cells. E633A and E597A/H602A variants exhibited reduction in both activities by 50–60% compared to recombinant wild-type ceruloplasmin. The variant E935A/H940A had reduced ferroxidase activity (50%) but unaltered diamine oxidase activity, whereas the variant E971A exhibited enhanced diamine oxidase activity. For the L329M variant, both activities were identical to those of wild-type ceruloplasmin. 相似文献
3.
Bruce X. Wong Scott Ayton Linh Q. Lam Peng Lei Paul A. Adlard Ashley I. Bush James A. Duce 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014
Background
Iron oxidation is thought to be predominantly handled enzymatically in the body, to minimize spontaneous combustion with oxygen and to facilitate cellular iron export by loading transferrin. This process may be impaired in disease, and requires more accurate analytical assays to interrogate enzymatic- and auto-oxidation within a physiologically relevant environment.Method
A new triplex ferroxidase activity assay has been developed that overcomes the previous assay limitations of measuring iron oxidation at a physiologically relevant pH and salinity.Results
Revised enzymatic kinetics for ceruloplasmin (Vmax ≈ 35 μM Fe3 +/min/μM; Km ≈ 15 μM) are provided under physiological conditions, and inhibition by sodium azide (Ki for Ferric Gain 78.3 μM, Ki for transferrin loading 8.1 × 104 μM) is quantified. We also used this assay to characterize the non-enzymatic oxidation of iron that proceeded linearly under physiological conditions.Conclusions and general significance
These findings indicate that the requirement of an enzyme to oxidize iron may only be necessary under conditions of adverse pH or anionic strength, for example from hypoxia. In a normal physiological environment, Fe3 + incorporation into transferrin would be sufficiently enabled by the biological polyanions that are prevalent within extracellular fluids. 相似文献4.
V. N. Zaitsev I. Zaitseva M. Papiz P. F. Lindley 《Journal of biological inorganic chemistry》1999,4(5):579-587
Ceruloplasmin is a multi-copper oxidase, which contains most of the copper present in the plasma. It is an acute-phase reactant that exhibits a two- to three-fold increase over the normal concentration of 300?μg/ml in adult plasma. However, the precise physiological role(s) of ceruloplasmin has been the subject of intensive debate and it is likely that the enzyme has a multi-functional role, including iron oxidase activity and the oxidation of biogenic amines. The three-dimensional X-ray structure of the human enzyme was elucidated in 1996 and showed that the molecule was composed of six cupredoxin-type domains arranged in a triangular array. There are six integral copper atoms per molecule (mononuclear sites in domains 2, 4 and 6 and a trinuclear site between domains 1 and 6) and two labile sites with roughly 50% occupancy. Further structural studies on the binding of metal cations by the enzyme indicated a putative mechanism for ferroxidase activity. In this paper we report medium-resolution X-ray studies (3.0–3.5?Å) which locate the binding sites for an inhibitor (azide) and various substrates [aromatic diamines, biogenic amines and (+)-lysergic acid diethylamide, LSD]. The binding site of the azide moiety is topologically equivalent to one of the sites reported for ascorbate oxidase. However, there are two distinct binding sites for amine substrates: aromatic diamines bind on the bottom of domain 4 remote from the mononuclear copper site, whereas the biogenic amine series typified by serotonin, epinephrine and dopa bind in close vicinity to that utilised by cations in domain 6 and close to the mononuclear copper. These binding sites are discussed in terms of possible oxidative mechanisms. The binding site for LSD is also reported. 相似文献
5.
The Fet3 protein in Saccharomyces cerevisiae and mammalian ceruloplasmin are multicopper oxidases (MCO) that are required for iron homeostasis via their catalysis of the ferroxidase reaction, 4Fe(2+)+O(2)+4H(+)-->4Fe(3+)+2H(2)O. The enzymes may play an essential role in copper homeostasis since they exhibit a strikingly similar kinetic activity towards Cu(1+) as substrate. In contrast, laccase, an MCO that exhibits weak activity towards Fe(2+), exhibits a similarly weak activity towards Cu(1+). Kinetic analyses of the Fet3p reaction demonstrate that the ferroxidase and cuprous oxidase activities are due to the same electron transfer site on the enzyme. These two ferroxidases are fully competent kinetically to play a major role in maintaining the cuprous-cupric redox balance in aerobic organisms. 相似文献
6.
Ceruloplasmin (CP) oxidises low density lipoprotein (LDL). The oxidising potential depends on the formation of Cu(+)-CP which is redox-cycled during oxidation. Homocysteine (HCY) reduces free Cu(2+), potentiating its cell-damaging property. We show that HCY enhanced LDL oxidation by CP, but did not activate the LDL oxidising potential of Cu(2+)-diamine oxidase. Selective removal of the redox-active Cu(2+) abolished the LDL oxidase activity of CP. However, HCY partially restored the LDL oxidase activity of redox-copper depleted CP, indicating that the remaining six copper atoms in CP may also be involved in the process. Spectroscopic and oxidation inhibition studies using the Cu(+)-reagent bathocuproine revealed that HCY induced Cu(+)-CP formation, thus promoting its LDL oxidase activity. 相似文献
7.
Rolf Arthur L¿vstad 《Biometals》1997,10(2):123-126
The ferroxidase activity of ceruloplasmin is often determined according to the method of Johnson et al. (1967), using apotransferrin for trapping ferric ions generated by the enzyme; spectrophotometrically monitoring the Fe–transferrin formation at pH6.0. Reports have shown that ascorbate inhibits this reaction, and it is hypothesized that the effect could be of physiological significance in individuals with a high ascorbate to ceruloplasmin ratio in plasma (e.g. premature babies).The present study shows that the inhibitory effect of ascorbate rapidly decreases with increasing pH. At pH7.4 no significant effect was observed, the result suggesting that ascorbate is not a physiological inhibitor of ceruloplasmin. Furthermore, experiments demonstrate that at acidic pH the inhibitory effect of ascorbate on the rate of Fe–transferrin formation is not primarily due to an interaction with ceruloplasmin, but to a reduction of enzymically generated ferric ions before they are bound to apotransferrin. 相似文献
8.
J. Fetter Jonathan Cohen D. Danger Joann Sanders-Loehr E. C. Theil 《Journal of biological inorganic chemistry》1997,2(5):652-661
Ferritins uniquely direct the vectorial transfer of hydrated Fe(II)/Fe(III) ions to a condensed ferric phase in the central cavity of the soluble protein. Secondary, tertiary and quaternary structure are conserved in ferritin, but only five amino acid residues are conserved among all known ferritins. The sensitivity of ferroxidation rates to small differences in primary sequence between ferritin subunits that are cell-specifically expressed or to the conservative replacement of the conserved tyrosine 30 residue was demonstrated by examining recombinant (frog) H-type (red blood cell predominant) and M-type subunit (liver predominant) proteins which are both fast ferritins; the proteins form two differently colored Fe(III)-protein complexes absorbing at 550?nm or 650?nm, respectively. The complexes are convenient reporters of Fe(III)-protein interaction because they are transient in contrast to the Fe(III)-oxy complexes measured in the past at 310–420?nm, which are stable because of contributions from the mineral itself. The A650-nm species formed 18-fold faster in the M-subunit protein than did the 550-nm species in H-subunit ferritin, even though all the ferroxidase residues are the same; the Vmax was fivefold faster but the Hill coefficents were identical (1.6), suggesting similar mechanisms. In H-subunit ferritin, substitution of phenylalanine for conserved tyrosine 30 (located in the core of the subunit four-helix bundle) slowed ferroxidation tenfold, whereas changing surface tyrosine 25 or tyrosine 28 had no effect. The Fe(III)-tyrosinate was fortunately not changed by the mutation, based on the resonance Raman spectrum, and remained a suitable reporter for Fe(III)-protein interactions. Thus, the A550/650?nm can also report on post-oxidation events such as transport through the protein. The impact of Y30F on rates of formation of Fe(III)-protein complexes in ferritin, combined with Mössbauer spectroscopic studies that showed the parallel formation of multiple Fe(III) postoxidation species (three dinuclear oxy and one trinuclear oxy species) (A. S. Periera et al., Biochemistry 36?:?7917–7927, 1997) and the loss of several of the multimeric Fe(III) post-oxidation species in a Y30F alteration of human recombinant H-ferritin (E. R. Bauminger et al., Biochem J. 296?:?709–719, 1993), indicate that at least one of the pathways for Fe oxidation/transfer in ferritin is through the center of the four-helix bundle and is influenced by structural features dependent on tyrosine 30. 相似文献
9.
BackgroundCeruloplasmin (Cp) is a major copper-binding protein produced in the liver and delivers copper to extrahepatic organs. Patients with myocardial infarction are often featured by an elevation of serum copper concentrations due to copper efflux from ischemic hearts. The present study was undertaken to test the hypothesis that serum copper elevation leads to up-regulation of hepatic Cp in myocardial infarction.MethodsAdult male Sprague-Dawley rats were subjected to left anterior descending (LAD) coronary artery ligation to induce myocardial infarction. Serum copper and Cp levels, as well as changes in hepatic Cp and copper-transporting P-type ATPase (Atp7b), were determined from blood and liver samples collected on day 1, 4, or 7 after the operation.ResultsSerum copper concentrations were significantly increased on day 4 after LAD ligation, accompanied by an increase in serum Cp levels and activities. Concomitantly, the protein levels of Cp and copper exporter, Atp7b, were also significantly increased in the liver. Furthermore, inhibiting the increase of serum copper by a copper chelator, triethylenetetramine (TETA), effectively abolished the elevated Cp activity after LAD ligation.ConclusionThese results indicate that serum Cp elevation in response to myocardial ischemia most likely resulted from the increased hepatic Cp production, which in turn was more responsive to serum copper elevation than inflammatory response following myocardial ischemia. 相似文献
10.
Human ceruloplasmin (CP) is a multifunctional copper-binding protein produced in the liver. CP oxidizes Fe2+ to Fe3+, decreasing the concentration of Fe2+ available for generating harmful oxidant species. CP is also a potent inhibitor of leukocyte myeloperoxidase (MPO) (Kd=130 nM), a major source of oxidants in vivo. Rheumatoid arthritis (RA) is an inflammatory autoimmune disease affecting flexible joints and characterized by activation of both inflammatory and coagulation processes. Indeed, the levels of CP, MPO, and thrombin are markedly increased in the synovial fluid of RA patients. Here we show that thrombin cleaves CP in vitro at 481Arg–Ser482 and 887Lys–Val888 bonds, generating a nicked species that retains the native-like fold and the ferroxidase activity of the intact protein, whereas the MPO inhibitory function of CP is abrogated. Analysis of the synovial fluid of 24 RA patients reveals that CP is proteolytically degraded to a variable extent, with a fragmentation pattern similar to that observed with thrombin in vitro, and that proteolysis is blocked by hirudin, a highly potent and specific thrombin inhibitor. Using independent biophysical techniques, we show that thrombin has intrinsic affinity for CP (Kd=60–270 nM), independent of proteolysis, and inhibits CP ferroxidase activity (KI=220±20 nM). Mapping of thrombin binding sites with specific exosite-directed ligands (i.e., hirugen, fibrinogen γ′-peptide) and thrombin analogues having the exosites variably compromised (i.e., prothrombin, prethrombin-2, βT-thrombin) reveals that the positively charged exosite-II of thrombin binds to the negatively charged upper region of CP, while the protease active site and exosite-I remain accessible. These results suggest that thrombin can exacerbate inflammation in RA by impairing the MPO inhibitory function of CP via proteolysis and by competitively inhibiting CP ferroxidase activity.Notably, local administration of hirudin, a highly potent and specifc thrombin inhibitor, reduces the concentration of active MPO in the synovial fluid of RA patients and has a beneficial effect on the clinical symptoms of the disease. 相似文献
11.
Timothy P. Ryan Dennis M. Miller Steven D. Aust 《Journal of biochemical and molecular toxicology》1993,8(1):33-39
The effects of transition metals on nonenzymatic and ceruloplasmin catalyzed epinephrine oxidation were investigated by studying rates of epinephrine oxidation in purified buffers and in the presence of metal chelating agents. We found that epinephrine does not “autoxidize” in sodium chloride solutions prepared with deionized water that was further purified by chromatography over Chelex 100 resin prior to use. Epinephrine was oxidized rapidly in sodium chloride prepared with tap water (1.20±0.12 nmoles/min) or in deionized water (0.40±0.80 nmoles/min), but this oxidation was prevented by the addition of Desferal, a potent metal chelating agent. Epinephrine oxidation was enhanced upon the addition of ceruloplasmin, and this oxidation rate could be slowed, but not eliminated, by the addition of Desferal. If epinephrine solutions were preincubated for 72 hours with Desferal prior to ceruloplasmin addition, however, no oxidation was observed. Epinephrine was shown to form colored complexes with both iron and copper at pH 7.0. The Fe(III)-epinephrine complex was much more stable than was the Cu(II)-epinephrine complex. Oxygen consumption studies of ceruloplasmin catalyzed epinephrine oxidation showed that copper was a better promoter of epinephrine oxidation than was iron, suggesting that ceruloplasmin-catalyzed epinephrine oxidation results from adventitious copper bound to the purified enzyme. In light of these results, the physiological relevance of ceruloplasmin catalyzed oxidation of biogenic amines may be minor. 相似文献
12.
We demonstrated previously that loading iron into ferritin via its own ferroxidase activity resulted in damage to the ferritin
while ferritin loaded by ceruloplasmin, a copper-containing ferroxidase, was not damaged and had similar characteristics to
native ferritin (Welch et al. (2001) Free Radic Biol Med 31:999–1006). Interestingly, it has been suggested that the formation of hemosiderin, a proposed degradation
product of ferritin, is increased in animals deficient in copper. In this study, groups of rats were fed normal diets, copper
deficient diets, iron supplemented diets, or copper deficient-iron supplemented diets for 60 days. Rats fed copper-deficient
diets had no detectable active serum ceruloplasmin, which indicates that they were functionally copper deficient. There was
a significant increase in the amount of iron in isolated hemosiderin fractions from the livers of copper-deficient rats, even
more than that found in rats fed only an iron-supplemented diet. Histological analysis showed that copper-deficient rats had
iron deposits (which are indicative of hemosiderin) in their hepatocytes and Kupffer cells, whereas rats fed diets sufficient
in copper only had iron deposits in their Kupffer cells. Histologic evidence of iron deposition was more pronounced in rats
fed diets that were deficient in copper. Additionally, sucrose density-gradient sedimentation profiles of ferritin loaded
with iron in vitro via its own ferroxidase activity was found to have similarities to that of the sedimentation profile of
the hemosiderin fraction from rat livers. The implications of these data for the possible mechanism of hemosiderin formation
are discussed. 相似文献
13.
Musci G Polticelli F Bonaccorsi di Patti MC 《World journal of biological chemistry》2014,5(2):204-215
Safe trafficking of iron across the cell membrane is a delicate process that requires specific protein carriers. While many proteins involved in iron uptake by cells are known, only one cellular iron export protein has been identified in mammals: ferroportin(SLC40A1). Ceruloplasmin is a multicopper enzyme endowed with ferroxidase activity that is found as a soluble isoform in plasma or as a membrane-associated isoform in specific cell types. According to the currently accepted view, ferrous iron transported out of the cell by ferroportin would be safely oxidized by ceruloplasmin to facilitate loading on transferrin. Therefore, the ceruloplasminferroportin system represents the main pathway for cellular iron egress and it is responsible for physiological regulation of cellular iron levels. The most recent findings regarding the structural and functional features of ceruloplasmin and ferroportin and their relationship will be described in this review. 相似文献
14.
Phloem transport of amino acids in two Brassica napus L. genotypes and one B. carinata genotype in relation to their seed protein content 总被引:2,自引:0,他引:2
In order to investigate the relationship between the amino acid concentration in the phloem sap of leaves and the protein
content in seeds, two Brassica napus genotypes and one B. carinata genotype with low, medium and high seed protein contents were analyzed. Phloem sap was collected from the B. napus winter rapeseed breeding line DSV15 with 19% protein of dry weight in the seeds, the spring cultivar ‘Duplo’ with 25% protein
in the seeds and from the B. carinata line BRA1151/90 with 39% protein in the seeds by using the aphid-stylet technique. The total amino acid contents measured
in the phloem varied considerably among the three genotypes analysed, and correlated positively with their respective seed
protein contents. The total amino acid-to-sucrose ratio was lowest in B. napus line DSV15 which had the lowest seed protein content and highest in the B. carinata line BRA1151/90 which had the highest seed protein content. The amino-N translocation in the phloem during the light period
was about 2-fold higher in the B. carinata line BRA1151/90 than in the B. napus lines Dulpo and DSV15. Predominant amino acids in the phloem were glutamine and glutamate, followed by serine, aspartate,
and threonine. The amino acid patterns in the leaves resembled those in the phloem, although their absolute concentrations
were higher in the phloem than in the cytosol of mesophyll tissue. Furthermore, the concentration gradient of amino acids
between the cytosol of mesophyll cells and the phloem was higher in the B. carinata line BRA1151/90 than in the B. napus lines Duplo and DSV15. These results lead to the conclusion that the phloem translocation of amino-N and the phloem loading
process of amino acids are decisive factors for the protein content in the seeds of Brassica species.
Received: 28 November 1999 / Accepted: 10 April 2000 相似文献
15.
M. Giorgetti I. Ascone M. Berrettoni P. Conti S. Zamponi R. Marassi 《Journal of biological inorganic chemistry》2000,5(2):156-166
An in situ X-ray absorption spectroscopy (XAS) spectroelectrochemical study of aquocobalamin (system B12a-B12r-B12s) has been carried out in aqueous solutions buffered at different pH values. To the best of our knowledge, this is the first
structural study of aquocobalamin at room temperature under controlled oxidation conditions. Most of the previous work was
in fact performed using frozen samples chemically treated to produce the species. The spectroelectrochemical approach offers
several advantages: (1) the reduction products may be studied without poisoning the system with chemical reductive reagents
and (2) any possible variation of the oxidation state owing to the electrons produced by the incident beam is avoided as the
electrode, under potentiostatic control, acts as a scavenger. The spectroelectrochemical approach, together with more careful
data analysis, has led to an improved interpretation of the XAS data. These conditions were not met in previous works where
the oxidation state was not controlled and multiple scattering contributions were not taken into account. The general shape
of the XAS spectra of the different species is not greatly affected by pH. A signature for the base-off square-planar coordination
has been evidenced for the Co(II) compound at basic pH. A new signature for Co(I), indicating square-planar coordination,
has been identified on the experimental spectra and simulated in theoretical X-ray absorption near-edge structure (XANES)
studies. The flexibility of the electrochemical approach, that permits to unambiguously establish the formal oxidation state,
has led to very reliable values for energy shift and peak intensity variations. The experimental XANES and extended X-ray
absorption fine structure (EXAFS) spectra with a very good signal-to-noise ratio have been processed using the GNXAS package
that takes into account multiple scattering contributions. EXAFS and XANES independent analysis result in the same structural
model. The reduction from Co(III) to Co(II) produces the most significant structural changes: the cobalt coordination number
decreases from six to five, and the edge position shifts by 2.4±0.3 eV. In addition, the XANES spectra are strongly modified.
The reduction from Co(II) to Co(I) produces mainly electronic effects with no apparent change of the coordination number.
A discussion of the limits and potentialities of EXAFS in this type of study has also been included.
Received: 26 July 1999 / Accepted: 22 October 1999 相似文献
16.
G. Besnard Y. Griveau M. C. Quillet H. Serieys P. Lambert D. Vares A. Bervillé 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1997,94(1):131-138
A method based upon targetting of intro-gressed markers in a Phomopsis-resistant line (R) of cultivated sunflower, issuing
from a H. argophyllus cross was used to mark the Phomopsis resistance regions. Our study was based upon 203 families derived from a cross between an inbred line susceptible to Phomopsis (S1) and the introgressed resistant line (R).
Families were checked for Phomopsis resistance level in a design with replicated plots and natural infection was re-inforced
by pieces of contaminated stems. Thirty four primers were employed for RAPD analysis. Out of 102 polymorphic fragments between
(S1) and H. argophyllus, seven were still present in (R) suggesting that they marked introgressions of H. argophyllus into (R). The plants were scored for the presence or absence of 19 fragments obtained from five primers, and the relationships between
the presence/absence of fragments in plants and Phomopsis resistance/susceptiblity in the progenies was determined by using an analysis of variance. We found that at least two introgressed regions, as well as favourable
factors from sunflower, contributed to the level of Phomopsis resistance in cultivated sunflower.
Received: 28 June 1996 / Accepted: 5 July 1996 相似文献
17.
Genetic engineering approaches to improve the bioavailability and the level of iron in rice grains 总被引:28,自引:0,他引:28
P. Lucca R. Hurrell I. Potrykus 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2001,102(2-3):392-397
Iron deficiency is the most widespread micronutrient deficiency world-wide. A major cause is the poor absorption of iron from
cereal and legume-based diets high in phytic acid. We have explored three approaches for increasing the amount of iron absorbed
from rice-based meals. We first introduced a ferritin gene from Phaseolus vulgaris into rice grains, increasing their iron content up to two-fold. To increase iron bioavailability, we introduced a thermotolerant
phytase from Aspergillus fumigatus into the rice endosperm. In addition, as cysteine peptides are considered a major enhancer of iron absorption, we overexpressed
the endogenous cysteine-rich metallothionein-like protein. The content of cysteine residues increased about seven-fold and
the phytase level in the grains about 130-fold, giving a phytase activity sufficient to completely degrade phytic acid in
a simulated digestion experiment. High phytase rice, with an increased iron content and rich in cysteine-peptide, has the
potential to greatly improve iron nutrition in rice-eating populations.
Received: 15 April 2000 / Accepted: 12 May 2000 相似文献
18.
RFLP mapping of resistance to chlorosis induction by Pyrenophora tritici-repentis in wheat 总被引:2,自引:0,他引:2
J. D. Faris J. A. Anderson L. J. Francl J. G. Jordahl 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1997,94(1):98-103
Tan spot, caused by Pyrenophora tritici-repentis, is an economically important disease in major wheat production areas. The fungus can produce two genetically distinct symptoms
on leaves of susceptible wheat genotypes: tan necrosis (nec) and extensive chlorosis (chl). Our objectives were to determine
the number of genes conditioning resistance to tan spot in a population of wheat recombinant inbred lines, and map the chromosomal
location of the resistance genes using RFLPs. Conidia produced by the P. tritici-repentis isolate Pti2 (nec+chl+) were used to inoculate seedlings of 135 recombinant inbred lines derived from the cross of the synthetic
hexaploid wheat W-7984 with Opata 85. A subset of the population was inoculated with conidia produced by the isolates D308
(nec−chl+) and 86-124 (nec+chl−). Inoculated seedlings were rated on a scale of 1 to 5 based on lesion type. Necrosis-inducing
culture filtrate produced by the isolate 86-124 was also used to screen the entire population. A map consisting of 532 markers
was employed to identify significant associations between marker loci and tan spot resistance. The entire population was insensitive
to culture filtrate produced by the isolate 86-124, and the entire subset was resistant to conidial inoculation of the same
isolate. The population segregated for reaction to isolates D308 and Pti2, indicating that this population segregates for
resistance to extensive chlorosis only, and not to tan necrosis. RFLP analysis indicated the presence of a gene with a major
effect in 1AS, a gene with a minor effect in 4AL, and an interaction between the 1AS gene and a gene in 2DL. Together, these
loci explained 49.0% of the variation in this population for resistance to tan spot produced by the isolate Pti2. Two regions
one in 1BL and one in 3BL, were significantly associated with resistance to extensive chlorosis, but were not significant
in the multiple regression model. It should be feasible to introgress these resistance loci into adapted genetic backgrounds
by using a marker-assisted selection scheme.
Received: 30 March 1996 / Accepted: 31 May 1996 相似文献
19.
Previous studies have demonstrated that 2-hydroxy-1-naphthaldehyde isonicotinoyl hydrazone (NIH) and several other aroylhydrazone
chelators possess anti-neoplastic activity due to their ability to bind intracellular iron. In this study we have examined
the structure and properties of NIH and its FeIII complex in order to obtain further insight into its anti-tumour activity. Two tridentate NIH ligands deprotonate upon coordination
to FeIII in a meridional fashion to form a distorted octahedral, high-spin complex. Solution electrochemistry of [Fe(NIH–H)2]+ shows that the trivalent oxidation state is dominant over a wide potential range and that the FeII analogue is not a stable form of this complex. The fact that [Fe(NIH–H)2]+ cannot cycle between the FeII and FeIII states suggests that the production of toxic free-radical species, e.g. OH
.
or O2
.
–, is not part of this ligand's cytotoxic action. This suggestion is supported by cell culture experiments demonstrating that
the addition of FeIII to NIH prevents its anti-proliferative effect. The chemistry of this chelator and its FeIII complex are discussed in the context of understanding its anti-tumour activity.
Received: 12 November 1998 / Accepted: 9 February 1999 相似文献
20.
Molecular genetics of the y locus in pepper: its relation to capsanthin-capsorubin synthase and to fruit color 总被引:2,自引:0,他引:2
S. Popovsky I. Paran 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2000,101(1-2):86-89
Classical genetic studies have determined that the yellow fruit color in pepper is recessive to red in the locus y. We studied the relation of the y locus with the gene coding for capsanthin-capsorubin synthase (CCS) that synthesizes the red carotenoid pigments in the mature
fruit. Cosegregation of y and CCS in populations derived from crosses between plants bearing red×white and red×yellow fruits indicated the correspondence
of the two genes. We obtained indications for the occurrence of a deletion in the CCS gene in plants containing the recessive
y allele. This deletion did not contain the distal 220 bp of the 3′ end of the gene. We used the CCS gene to determine the
genotype of peppers with different fruit colors at the y locus. In BC1 segregants from a red×white cross, the red and peach-fruited progenies had the wild-type allele at the CCS locus, while the
orange, yellow and white-fruited progenies had the mutant allele. Screening orange-fruited cultivars with CCS as well as segregation
analysis of CCS in an additional red×white cross indicated two possible genotypes of the orange fruit color in this locus.
Received: 25 January 1999 / Accepted: 16 August 1999 相似文献