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排序方式: 共有540条查询结果,搜索用时 62 毫秒
31.
Characterization of activity and expression of isocitrate lyase in Mycobacterium avium and Mycobacterium tuberculosis 总被引:5,自引:0,他引:5 下载免费PDF全文
Analysis by two-dimensional gel electrophoresis revealed that Mycobacterium avium expresses several proteins unique to an intracellular infection. One abundant protein with an apparent molecular mass of 50 kDa was isolated, and the N-terminal sequence was determined. It matches a sequence in the M. tuberculosis database (Sanger) with similarity to the enzyme isocitrate lyase of both Corynebacterium glutamicum and Rhodococcus fascians. Only marginal similarity was observed between this open reading frame (ORF) (termed icl) and a second distinct ORF (named aceA) which exhibits a low similarity to other isocitrate lyases. Both ORFs can be found as distinct genes in the various mycobacterial databases recently published. Isocitrate lyase is a key enzyme in the glyoxylate cycle and is essential as an anapleurotic enzyme for growth on acetate and certain fatty acids as carbon source. In this study we express and purify Icl, as well as AceA proteins, and show that both exhibit isocitrate lyase activity. Various known inhibitors for isocitrate lyase were effective. Furthermore, we present evidence that in both M. avium and M. tuberculosis the production and activity of the isocitrate lyase is enhanced under minimal growth conditions when supplemented with acetate or palmitate. 相似文献
32.
In the Clostridium beijerinckii flavodoxin, the reduction of the flavin mononucleotide (FMN) cofactor is accompanied by a local conformation change in which the Gly57-Asp58 peptide bond "flips" from primarily the unusual cis O-down conformation in the oxidized state to the trans O-up conformation such that a new hydrogen bond can be formed between the carbonyl group of Gly57 and the proton on N(5) of the neutral FMN semiquinone radical [Ludwig, M. L., Pattridge, K. A., Metzger, A. L., Dixon, M. M., Eren, M., Feng, Y., and Swenson, R. P. (1997) Biochemistry 36, 1259-1280]. This interaction is thought to contribute to the relative stabilization of the flavin semiquinone and may be at least partially responsible for the substantial separation of the midpoint potentials of the two one-electron reduction steps. Through a series of amino acid substitutions, the above cited study demonstrated the critical role of the often conserved glycine residue in this process. However, it has not been directly established experimentally as to whether these substitutions brought about the changes in the midpoint potentials by altering the strength of this hydrogen-bonding interaction as proposed. In this study, the relative strengths of the FMN N(5)H.O57 hydrogen bond in wild type and the G57A, G57N, and G57T mutants were evaluated by measuring the temperature dependency of the chemical shift for the proton on N(5) of the fully reduced cofactor by 1H-15N HSQC nuclear magnetic resonance spectroscopy. Based on the established correlation between the temperature coefficient of amide protons and the strength of hydrogen bonding in small peptides, the apparent strength of the N(5)H.O57 interaction was found to depend on the properties of the side chain at position 57. The glycine residue found in the wild-type flavodoxin appears to provide the strongest interaction while the beta-branched side chain in the G57T mutant provides the weakest. A good correlation was noted between the temperature coefficients of N(5)H and the one-electron reduction potential for the ox/sq couple as well as the binding free energy of the FMN semiquinone in this group of mutants. These results provide more direct quantitative evidence that support the previous hypothesis that this conformation change and the associated formation of the hydrogen bonding interaction with N(5)H of the reduced FMN represent an important means of stabilizing the neutral semiquinone and in modulating the oxidation-reduction potentials of the flavin cofactor in this and perhaps other flavodoxins. 相似文献
33.
Swenson DL Warfield KL Kuehl K Larsen T Hevey MC Schmaljohn A Bavari S Aman MJ 《FEMS immunology and medical microbiology》2004,40(1):27-31
Marburg virus (MARV), the causative agent of a severe hemorrhagic fever, has a characteristic filamentous morphology. Here we report that co-expression of MARV glycoprotein and matrix protein (VP40) in mammalian cells leads to spontaneous budding of filamentous particles strikingly similar to wild-type MARV. In addition, these particles elicit an immune response in BALB/c mice. The generation of non-replicating Marburg virus-like particles (VLPs) should significantly facilitate the research on molecular mechanisms of MARV assembly and release. Furthermore, VLPs may be an excellent vaccine candidate against Marburg infection. 相似文献
34.
Liu C Gaça MD Swenson ES Vellucci VF Reiss M Wells RG 《The Journal of biological chemistry》2003,278(13):11721-11728
Hepatic stellate cells are the primary cell type responsible for matrix deposition in liver fibrosis, undergoing a process of transdifferentiation into fibrogenic myofibroblasts. These cells, which undergo a similar transdifferentiation process when cultured in vitro, are a major target of the profibrogenic agent transforming growth factor-beta (TGF-beta). We have studied activation of the TGF-beta downstream signaling molecules Smads 2, 3, and 4 in hepatic stellate cells (HSC) cultured in vitro for 1, 4, and 7 days, with quiescent, intermediate, and fully transdifferentiated phenotypes, respectively. Total levels of Smad4, common to multiple TGF-beta superfamily signaling pathways, do not change as HSC transdifferentiate, and the protein is found in both nucleus and cytoplasm, independent of treatment with TGF-beta or the nuclear export inhibitor leptomycin B. TGF-beta mediates activation of Smad2 primarily in early cultured cells and that of Smad3 primarily in transdifferentiated cells. The linker protein SARA, which is required for Smad2 signaling, disappears with transdifferentiation. Additionally, day 7 cells demonstrate constitutive phosphorylation and nuclear localization of Smad 2, which is not affected by pretreatment with TGF-beta-neutralizing antibodies, a type I TGF-beta receptor kinase inhibitor, or activin-neutralizing antibodies. These results demonstrate essential differences between TGF-beta-mediated signaling pathways in quiescent and in vitro transdifferentiated hepatic stellate cells. 相似文献
35.
A new identity for MLK3 as an NIMA-related,cell cycle-regulated kinase that is localized near centrosomes and influences microtubule organization 总被引:1,自引:0,他引:1 下载免费PDF全文
Although conserved counterparts for most proteins involved in the G(2)/M transition of the cell cycle have been found in all eukaryotes, a notable exception is the essential but functionally enigmatic fungal kinase NIMA. While a number of vertebrate kinases have been identified with catalytic domain homology to NIMA, none of these resemble NIMA within its extensive noncatalytic region, a region critical for NIMA function in Aspergillus nidulans. We used a bioinformatics approach to search for proteins with homology to the noncatalytic region of NIMA and identified mixed lineage kinase 3 (MLK3). MLK3 has been proposed to serve as a component in MAP kinase cascades, particularly those resulting in the activation of the c-Jun N-terminal kinase (JNK). Here we describe the first in-depth study of endogenous MLK3 and report that, like NIMA, MLK3 phosphorylation and activity are enhanced during G(2)/M, whereas JNK remains inactive. Coincident with the G(2)/M transition, a period marked by dramatic reorganization of the cytoplasmic microtubule network, endogenous MLK3 transiently disperses away from the centrosome and centrosomal-proximal sites where it is localized during interphase. Furthermore, when overexpressed, MLK3, like NIMA, localizes to the centrosomal region, induces profound disruption of cytoplasmic microtubules and a nuclear distortion phenotype that differs from mitotic chromosome condensation. Cellular depletion of MLK3 protein using siRNA technology results in an increased sensitivity to the microtubule-stabilizing agent taxol. Our studies suggest a new role for MLK3, separable from its function in the JNK pathway, that may contribute to promoting microtubule instability, a hallmark of M phase entry. 相似文献
36.
Netesova IG Swenson PD Osipova LP Kiselev NN Posukh OL Cherepanova NS Kazakovtseva MA Kashinskaia IuO Netesov SV 《Zhurnal mikrobiologii, epidemiologii, i immunobiologii》2001,(2):29-33
Blood samples taken from 231 native inhabitants of the village of Mendur-Sokkon located in the Republic of Altai (South-Western Siberia, Russia) were tested for the presence of virus hepatitis B (HBV) markers. 31 samples (13.4%) were found to contain HBsAg, 111 samples (48.05%) were found to contain total anti-HBc antibodies, 123 samples (53.24%) were found to contain anti-HBs antibodies and 15 blood samples (6.49%), anti-HBc antibodies without anti-HBs antibodies and HBsAg. The age-dependent distribution of the occurrence of HBV markers among the aboriginal population of the South Altal remained unchanged (69.9 +/- 7.9%) for the last 50 years. The vertical and horizontal routes of HBV transmissions were noted. The data obtained in this study are indicative of a highly endemic character of HBV of the territory of Mendur-Sokkon. HBsAg-positive blood samples were taken for HBsAg subtyping with the use of a panel of monoclonal antibodies. Two subtypes of HBsAg were detected: ayw1-2 and ayw3varB with the occurrence of 92.6% and 7.4%, i.e. distributed in the ratio 25/2. 相似文献
37.
Ohdan H Swenson KG Kruger Gray HS Yang YG Xu Y Thall AD Sykes M 《Journal of immunology (Baltimore, Md. : 1950)》2000,165(10):5518-5529
Human natural Abs against Galalpha1-3Galbeta1-4GlcNAc (Gal) epitopes are a major barrier to xenotransplantation. Studies in this report, which use combined multiparameter flow cytometric sorting and enzyme-linked immunospot assay, demonstrate that anti-Gal IgM-producing cells are found exclusively in a small B cell subpopulation (i.e., CD21(-/low) IgM(high) B220(low) CD5(-) Mac-1(-) 493(-) cells) in the spleens of alpha1, 3-galactosyltransferase-deficient mice. All IgM-producing cells were detected in a similar splenic subpopulation of alpha1, 3-galactosyltransferase-deficient and wild-type mice. A higher frequency of B cells with anti-Gal surface IgM receptors was observed in the peritoneal cavity than in the spleen, but these did not actively secrete Abs, and showed phenotypic properties of B-1b cells (CD21(-/low) IgM(high) CD5(-) CD43(+) Mac-1(+)). However, these became Mac-1(-) and developed anti-Gal Ab-producing activity after in vitro culture with LPS. The splenic B cells with anti-Gal receptors consisted of both Mac-1(+) B-1b cells and Mac-1(-) B-1b-like cells. The latter comprised most anti-Gal IgM-producing cells. Our studies indicate that anti-Gal natural IgM Abs are produced by a B1b-like, Mac-1(-) splenic B cell population and not by plasma cells or B-1a cells. They are consistent with a model whereby B-1b cells lose Mac-1 expression upon Ag exposure and that these, rather than plasma cells, become the major IgM Ab-producing cell population. 相似文献
38.
High rate of DNA loss in the Drosophila melanogaster and Drosophila virilis species groups 总被引:6,自引:3,他引:3
We recently proposed that patterns of evolution of non-LTR
retrotransposable elements can be used to study patterns of spontaneous
mutation. Transposition of non-LTR retrotransposable elements commonly
results in creation of 5' truncated, "dead-on-arrival" copies. These
inactive copies are effectively pseudogenes and, according to the neutral
theory, their molecular evolution ought to reflect rates and patterns of
spontaneous mutation. Maximum parsimony can be used to separate the
evolution of active lineages of a non-LTR element from the fate of the
"dead-on-arrival" insertions and to directly assess the relative
frequencies of different types of spontaneous mutations. We applied this
approach using a non-LTR element, Helena, in the Drosophila virilis group
and have demonstrated a surprisingly high incidence of large deletions and
the virtual absence of insertions. Based on these results, we suggested
that Drosophila in general may exhibit a high rate of spontaneous large
deletions and have hypothesized that such a high rate of DNA loss may help
to explain the puzzling dearth of bona fide pseudogenes in Drosophila. We
also speculated that variation in the rate of spontaneous deletion may
contribute to the divergence of genome size in different taxa by affecting
the amount of superfluous "junk" DNA such as, for example, pseudogenes or
long introns. In this paper, we extend our analysis to the D. melanogaster
subgroup, which last shared a common ancestor with the D. virilis group
approximately 40 MYA. In a different region of the same transposable
element, Helena, we demonstrate that inactive copies accumulate deletions
in species of the D. melanogaster subgroup at a rate very similar to that
of the D. virilis group. These results strongly suggest that the high rate
of DNA loss is a general feature of Drosophila and not a peculiar property
of a particular stretch of DNA in a particular species group.
相似文献
39.
40.
Paul J. Weldon Daryl J. Swenson John K. Olson William G. Brinkmeier 《Ethology : formerly Zeitschrift fur Tierpsychologie》1990,85(3):191-198
American alligators (Alligator mississippiensis) were offered raw meat or animal carcasses under water or on land in experiments where only chemoreception could be used to discriminate between these and control materials. Alligators also were presented with aqueous extracts and airborne chemicals from meat. Juvenile alligators, tested in indoor tanks, opened more cheesecloth packets containing meat than control packets when these materials were placed under water and on platforms above the water surface. Adult alligators, tested in outdoor semi-natural enclosures, removed more cheesecloth-wrapped meat presented under water, and more of both meat and raccoon (Procyon lotor) carcasses placed under perforated baskets on land, than control materials. Juvenile alligators, tested in tanks partially filled with water, exhibited more lateral head movements and mouth-openings to an aqueous beef extract than to water alone. Juvenile alligators, tested in an olfactometer, exhibited more gular pumps to airborne chemicals from beef than to distilled water. These experiments indicate that alligators may use chemical cues to locate food both on land and under water, and that they detect both water- and airborne chemicals from meat. 相似文献