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991.
Marc Yudkoff Yevgeny Daikhin Zhi-Ping Lin Liana Nissim Janet Stern David Pleasure Itzhak Nissim 《Journal of neurochemistry》1994,62(3):1192-1202
Abstract: The aim was to study the extent to which leu-cine furnishes α-NH2 groups for glutamate synthesis via branched-chain amino acid aminotransferase. The transfer of N from leucine to glutamate was determined by incubating astrocytes in a medium containing [15N]leucine and 15 unlabeled amino acids; isotopic abundance was measured with gas chromatography-mass spectrometry. The ratio of labeling in both [15N]glutamate/[15N]leucine and [2-15N]glutamine/[15N]leucine suggested that at least one-fifth of all glutamate N had been derived from leucine nitrogen. At the same time, enrichment in [15N]leucine declined, reflecting dilution of the 16N label by the unlabeled amino acids that were in the medium. Isotopic abundance in [16N]-isoleucine increased very quickly, suggesting the rapidity of transamination between these amino acids. The appearance of 15N in valine was more gradual. Measurement of branched-chain amino acid transaminase showed that the reaction from leucine to glutamate was approximately six times more active than from glutamate to leucine (8.72 vs. 1.46 nmol/min/mg of protein). However, when the medium was supplemented with α-ketoisocaproate (1 mM), the ketoacid of leucine, the reaction readily ran in the “reverse” direction and intraastrocytic [glutamate] was reduced by ~50% in only 5 min. Extracellular concentrations of α-ketoisocaproate as low as 0.05 mM significantly lowered intracellular [glutamate]. The relative efficiency of branched-chain amino acid transamination was studied by incubating astrocytes with 15 unlabeled amino acids (0.1 mM each) and [15N]glutamate. After 45 min, the most highly labeled amino acid was [15N]alanine, which was closely followed by [15N]leucine and [15N]isoleucine. Relatively little 15N was detected in any other amino acids, except for [15N]serine. The transamination of leucine was ~17 times greater than the rate of [1-14C]leucine oxidation. These data indicate that leucine is a major source of glutamate nitrogen. Conversely, reamination of a-ketoisocaproate, the ketoacid of leucine, affords a mechanism for the temporary “buffering” of intracellular glutamate. 相似文献
992.
Summary The P-43 ofEudiplodinium and homologous proteins in three other entodiniomorphid species, free-living ciliates, flagellates, and HeLa cells, were identified at the plasma membrane-cytoskeleton interface. Proteins cross-reacting with MAb B6 were also located at the ciliary inner surface of the plasma membrane. Due to the strong adhesion of the plasma membrane to the underlying cytoskeleton, classical extraction with detergents, urea, NaOH, and PTA, failed to separate the two components completely. However, the extraction properties of P-43, associated with its membrane-cytoskeleton interactive functions, suggest that this unglycosylated protein may present some analogies with proteins of the intermediate filaments. Their ubiquity and localization suggest that P-43 and MAb B6 crossreacting proteins may not be strictly epiplasmic but could be amphitropic proteins, strongly anchored to both the plasma membrane and the underlying microfilament framework, via protein-protein binding or by direct insertion in the lipid bilayer.Abbreviations BSA
bovine serum albumin
- Con A
concanavalin A
- EDTA
ethylene diamine tetraacetic acid
- EM
electron microscopy
- IF
intermediate filaments
- MAb
monoclonal antibody
- MET
2-mercaptoethanol
- MW
molecular weight
- PAb
polyclonal antibody
- PBS
phosphate buffered saline
- PMSF
phenylmethylsulfonyl fluoride
- PTA
phosphotungstic acid
- SDS
sodiumdodecyl sulfate
- TAME
Na-p-tosyl-arginine methyl ester
- TLCK
Na-p-tosyl-lysine chloromethyl ketone 相似文献
993.
A genomic clone containing the gl1–2 allele has been isolated and sequenced. The predicted amino acid sequence of the gl1–2 protein is identical to that of the GL1-Col allele up to position 201. At this point in the coding region of gl1–2 there is a deletion relative to the wild-type sequence that results in an in-frame stop codon at position 202. This deletion removes 27 amino acid residues, including a highly negatively charged region, from the predicted gl1–2 polypeptide. The loss of this negatively charged carboxy-terminal region from the gl1–2 product is most likely the cause of the partial loss of gene activity which results in a reduction in leaf trichome initiation. 相似文献
994.
Mare K. Saba-El-Leil Sylvain Rivard David Morse Mario Cappadocia 《Plant molecular biology》1994,24(4):571-583
A genomic clone of the S11 allele from the self-incompatibility locus (S locus) in Solanum chacoense Bitt. has been isolated by cross-hybridization to the S. chacoense S13 allele and sequenced. The sequence of the S11 allele contains all the features expected for S genes of the Solanaceae, and S11 expression, as assessed by northern blots and RNA-PCR, was similar to that of other S. chacoense S alleles. The S11 protein sequence shares 95% identity with the phenotypically distinct S13 protein of S. chacoense and is the gametophytic S allele with the highest similarity to an existing allele so far discovered. Only 10 amino acid changes differentiate the mature proteins from these two alleles, which sets a new lower limit to the number of changes that can produce an altered S allele specificity. The amino acid substitutions are not clustered, suggesting that an accumulation of random point mutations can generate S allele diversity. The S11 intron is unusual in that it could be translated in frame with the coding sequence, thus suggesting an additional mechanism for the generation of new S alleles. 相似文献
995.
Jesper L. Aamand Audrey H. Hobson Catherine M. Buckley Steen T. Jørgensen Borge Diderichsen David J. McConnell 《Molecular & general genetics : MGG》1994,245(5):556-564
An extracellular Pseudomonas cepacia lipase, LipA, is inactive when expressed in the absence of the product of the limA gene. Evidence has been presented that LimA is a molecular chaperone. The lipA and limA genes have been cloned in separate and independently inducible expression systems in Escherichia coli. These systems were used to test the molecular chaperone hypothesis by investigating whether LimA could activate presynthesized prelipase and whether presynthesized LimA could activate newly synthesized prelipase. The results show that LimA cannot activate presynthesized prelipase and that presynthesized LimA can activate only a limited number of de novo synthesized prelipase molecules. Co-immunoprecipitation of prelipase/lipase with LimA generated a 1:1 complex of prelipase/lipase and LimA. The results suggest that a 1:1 complex of LipA and LimA is required for prelipase processing and secretion of active lipase. 相似文献
996.
Steven R. Scofield David A. Jones Kate Harrison Jonathan D. G. Jones 《Molecular & general genetics : MGG》1994,244(2):189-196
Antibiotic resistance genes can act as either cell autonomous or non-cell autonomous genetic markers with which to monitor the excision of plant transposons. To convert spectinomycin resistance from a noncell autonomous resistance to cell autonomous resistance, a transit peptide for chloroplast localization from a petunia ribulose bisphosphate carboxylase (rbcS) gene was fused in-frame to the aadA gene, which confers spectinomycin and streptomycin resistance. Constructs were generated in which the expression of this chimeric gene was prevented by the presence, in the 5 untranslated leader, of the maize transposons Activator (Ac) or Dissociation (Ds). When progeny of tobacco or tomato plants transformed with these constructs were germinated on spectinomycin-containing medium, germinally revertant and somatically variegated individuals could be distinguished. 相似文献
997.
We describe a screen to isolate cDNAs encoding Drosophila mitosis inhibitors capable of suppressing the mitotic catastrophe phenotype resulting in Schizosaccharomyces pombe from the combination of the weel-50 mutation with either a deletion allele of mil1, or with overexpression of cdc25
+. One plasmid was isolated which could suppress the temperature sensitive lethality of both these strains. The cDNA in this plasmid encodes a protein highly homologous to the DEAD-box family of ATP-dependent RNA helicases, rather than to protein kinases as might be expected. It is possible that the RNA helicase described here may regulate entry into mitosis by down regulating the expression of other genes whose activity may be rate-limiting for entry into mitosis. 相似文献
998.
Peter R. Johnston Stephen J. Walker Jari A.K. Hyttinen David Kilpatrick 《Mathematical biosciences》1994,120(2)
The inverse problem of electrocardiography, the computation of epicardial potentials from body surface potentials, is influenced by the desired resolution on the epicardium, the number of recording points on the body surface, and the method of limiting the inversion process. To examine the role of these variables in the computation of the inverse transform, Tikhonov's zero-order regularization and singular value decomposition (SVD) have been used to invert the forward transfer matrix. The inverses have been compared in a data-independent manner using the resolution and the noise amplification as endpoints. Sets of 32, 50, 192, and 384 leads were chosen as sets of body surface data, and 26, 50, 74, and 98 regions were chosen to represent the epicardium.The resolution and noise were both improved by using a greater number of electrodes on the body surface. When 60% of the singular values are retained, the results show a trade-off between noise and resolution, with typical maximal epicardial noise levels of less than 0.5% of maximum epicardial potentials for 26 epicardial regions, 2.5% for 50 epicardial regions, 7.5% for 74 epicardial regions, and 50% for 98 epicardial regions. As the number of epicardial regions is increased, the regularization technique effectively fixes the noise amplification but markedly decreases the resolution, whereas SVD results in an increase in noise and a moderate decrease in resolution. Overall the regularization technique performs slightly better than SVD in the noise-resolution relationship.There is a region at the posterior of the heart that was poorly resolved regardless of the number of regions chosen. The variance of the resolution was such as to suggest the use of variable-size epicardial regions based on the resolution. 相似文献
999.
Changes in in vivo fluorescence quenching in rye and barley as a function of reduced PSII light harvesting antenna size 总被引:1,自引:0,他引:1
Stefan Falk Marianna Krol Denis P. Maxwell David A. Rezansoff Gordon R. Gray Norman P. A. Huner 《Physiologia plantarum》1994,91(4):551-558
The relationship between the size of the light harvesting antenna to photosystem II (LHCII) and quenching of non-photochemical and dark level fluorescence was studied in wild-type rye (Secale cereale L. cv. Musketeer) and barley (Hordeum vulgare L. cv. Gunilla) as well as in the barley chlorophyll b-less chlorina F2 mutant (H. vulgare L. cv. Dornaria, chlorina-F2). Exposure for 10 min to an irradiance of 500 μmol m?2 s?1 resulted in a strong (0.71–0.73) non-photochemical (qs) quenching of the fluorescence yield in wild-type (WT) material, while the barley chlorina F2-mutant was quenched to 75% of this level. Relaxation of qs in darkness revealed a fast initial decay, related to relaxation of the high-energy-state dependent (qE) part of qs. Etiolated seedlings of rye and barley exposed to intermittent light (IML) for 36 cycles of 2 min light and 118 min darkness had suppressed Chl b and LHCII-production in both WT rye and barley, while the barley chlorina F2-mutant became totally devoid of all LHCII-polypeptides. It was found that the levels of qs and qs were similar in control grown barley chlorina F2 and IML-grown WT rye and barley, but qs was reduced by 30 to 35% and qs by 50 to 65%, respectively, as compared to control-grown. WT plants. No significant qs could be detected in IML-grown barley chlorina F2. It is clear, from these changes in in vivo fluorescence quenching in rye and barley that a significant level of qs is detectable even in the absence of LHCII. Only when the proximal antennae are totally absent, does qE completely disappear. We conclude that the presence of LHCII is not an absolute requirement for qE-quenching and suggest that distal as well as proximal antenna may contribute to qE in vivo. 相似文献
1000.
We examined the effects of tail autotomy on survivorship and body growth of both adult and juvenile Uta stansburiana by directly manipulating tail condition. Tail loss decreased neither survivorship nor rate of body growth for individuals in two natural populations. Lack of an influence of tail loss on survivorship in these two populations may be the result of high mortality. Under high mortality any differential effects of tail loss will be lower than in populations facing lower mortality. Growth experiments in the laboratory demonstrated that, under conditions of minimal environmental variation and social interactions, there is no tradeoff between body growth and tail regeneration as has been suggested for other species of lizards. One possible reason for this difference is that U. stansburiana does not use the tail as a storage organ for lipids. The original and regenerated tails are composed mainly of protein. In general, any differential body growth between tailed and tailless individuals may be due to social interactions and not a diversion of limited energy into tail regeneration. 相似文献