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61.
Viral glycoproteins destined for apical or basolateral plasma membrane domains traverse the same Golgi apparatus during their intracellular transport in doubly infected Madin-Darby canine kidney cells 总被引:19,自引:32,他引:19
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M J Rindler I E Ivanov H Plesken E Rodriguez-Boulan D D Sabatini 《The Journal of cell biology》1984,98(4):1304-1319
Madin-Darby canine kidney (MDCK) cells can sustain double infection with pairs of viruses of opposite budding polarity (simian virus 5 [SV5] and vesicular stomatitis virus [VSV] or influenza and VSV), and we observed that in such cells the envelope glycoproteins of the two viruses are synthesized simultaneously and assembled into virions at their characteristic sites. Influenza and SV5 budded exclusively from the apical plasma membrane of the cells, while VSV emerged only from the basolateral surfaces. Immunoelectron microscopic examination of doubly infected MDCK cells showed that the influenza hemagglutinin (HA) and the VSV G glycoproteins traverse the same Golgi apparatus and even the same Golgi cisternae. This indicates that the pathways of the two proteins towards the plasma membrane do not diverge before passage through the Golgi apparatus and therefore that critical sorting steps must take place during or after passage of the glycoproteins through this organelle. After its passage through the Golgi, the HA accumulated primarily at the apical membrane, where influenza virion assembly occurred. A small fraction of HA did, however, appear on the lateral surface and was incorporated into the envelope of budding VSV virions. Although predominantly found on the basolateral surface, significant amounts of G protein were observed on the apical plasma membrane well before disruption of the tight junctions was detectable. Nevertheless, assembly of VSV virions was restricted to the basolateral domain and in doubly infected cells the G protein was only infrequently incorporated into the envelope of budding influenza virions. These observations indicate that the site of VSV budding is not determined exclusively by the presence of G polypeptides. Therefore, it is likely that, at least for VSV, other cellular or viral components are responsible for the selection of the appropriate budding domain. 相似文献
62.
Methionine aminopeptidase, known to be encoded by single genes in prokaryotes, is a cobalt-dependent enzyme that catalyzes the removal of N-terminal methionine residues from nascent polypeptides. Three ORFs encoding putative methionine aminopeptidases from the genome of cyanobacterium Synechocystis sp. strain PCC6803, designated as slr0786 (map-1), slr0918 (map-2) and sll0555 (map-3) were cloned and expressed in Escherichia coli. The purified recombinant proteins encoded by map-1 and map-3 had much higher methionine aminopeptidase activity than the recombinant protein encoded by map-2. Comparative analysis revealed that the three recombinant enzymes differed in their substrate specificity, divalent ion requirement, pH, and temperature optima. The broad activities of the iso-enzymes are discussed in light of the structural similarities with other peptidase families and their levels of specificity in the cell. Potential application of cyanobacterial MetAPs in the production of recombinant proteins used in medicine is proposed. This is the first report of a prokaryote harboring multiple methionine aminopeptidases.Abbreviations map Gene encoding methionine aminopeptidase - MetAP Methionine aminopeptidase - eMetAP-Ia Escherichia coli methionine aminopeptidase type Ia - yMetAP-Ib Yeast methionine aminopeptidase type Ib - yMetAP-IIa Yeast methionine aminopeptidase type IIa - hMetAP-IIb Human methionine aminopeptidase type IIb - pfMetAP–IIa Pyrococcus furiosis methionine aminopeptidase type Ia - bst MetAP-Ia Bacillus stearothermophilus methionine aminopeptidase type Ia - c1MetAP-Ia Cyanobacterial methionine aminopeptidase type Ia encoded by map-1 - c2MetAP-Ia Cyanobacterial methionine aminopeptidase type Ia encoded by map-2 - c3MetAP-Ib Cyanobacterial methionine aminopeptidase type Ib, ncoded by map-3 相似文献
63.
Spatial relationships among the transmembrane (TM) segments of alpha- and beta-subunits of the Na,K-ATPase molecule have been investigated using oxidative induction of disulfide bonds. The catalytic alpha-subunit contains 10 TM alpha-helices (H1-H10) with 9 Cys residues located within or close to the membrane moiety. There is one Cys residue in the single TM segment of beta-subunit (Hbeta). Previously, the cross-linking products containing the beta-subunit and two fragments of alpha-subunit (the N-terminal containing H1-H2 helices and the C-terminal containing H7-H10 helices) have been identified in experiments with membrane-bound or detergent-solubilized preparations of the membrane moiety of trypsin-digested Na,K-ATPase [Sarvazyan, N. A., Modyanov, N. N., and Askari, A. (1995) J. Biol. Chem. 270, 26528-26532 and Sarvazyan, N. A., Ivanov, A., Modyanov, N. N., and Askari, A. (1997) J. Biol. Chem. 272, 7855-7858]. Here, we have shown that Cu(2+)-phenanthroline treatment of digitonin-solubilized preparation provides the most efficient formation of intersubunit cross-linked product that is predominantly a dimer of beta-subunit and a 22-kDa C-terminal alpha-fragment containing H7-H10 helices. This cross-linked product was isolated and subjected to CNBr cleavage. The resulting fragments were electrophoretically separated and sequenced. A 17-kDa peptide composed of Ile853-Met942 alpha-fragment and Ala5-Met56 beta-fragment was identified as a product of intersubunit disulfide cross-link between Cys44 of Hbeta and either Cys911 or Cys930, located in H8. This provides the first direct experimental evidence of the juxtaposition of Hbeta and H8 within the Na,K-ATPase molecule. The second detected cross-linked product was composed of alpha-fragments Lys947-Met963 and Tyr974-Tyr1016 linked by induced disulfide bridge between Cys964 (H9) and Cys983 (H10). The spatial proximity of these Cys residues defines the mutual orientation of H9 and H10 helices of alpha-subunit. 相似文献
64.
K. A. Malsagova Yu. D. Ivanov T. O. Pleshakova A. F. Kozlov N. V. Krohin A. L. Kaysheva I. D. Shumov V. P. Popov O. V. Naumova B. I. Fomin D. A. Nasimov 《Biochemistry (Moscow) Supplemental Series B: Biomedical Chemistry》2014,8(3):220-225
The nanowire (NW) detection is one of the fast-acting and high-sensitive methods, which can recognize potentially relevant protein molecules. A NW-biosensor based on the silicon-on-insulator (SOI)-structures has been used for biospecific label-free real time detection of the NFATc1 (D-NFATc1) oncomarker. For this purpose, SOI-nanowires (NWs) were modified with aptamers against NFATc1 used as molecular probes. It was shown that using this biosensor it is possible to reach sensitivity of 10?15 M. This sensitivity was comparable to that of the NW-biosensor with immobilized antibodies used as macromolecular probes. The results demonstrate that approaches used in this study are promising for development of sensor elements for high-sensitive diagnostics of diseases. 相似文献
65.
Overcoming the Thermal Instability of Efficient Polymer Solar Cells by Employing Novel Fullerene‐Based Acceptors
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66.
Zhou Y Drummond DC Zou H Hayes ME Adams GP Kirpotin DB Marks JD 《Journal of molecular biology》2007,371(4):934-947
To determine the importance of single-chain Fv (scFv) affinity on binding, uptake, and cytotoxicity of tumor-targeting nanoparticles, the affinity of the epidermal growth factor receptor (EGFR) scFv antibody C10 was increased using molecular evolution and yeast display. A library containing scFv mutants was created by error-prone PCR, displayed on the surface of yeast, and higher affinity clones selected by fluorescence activated cell sorting. Ten mutant scFv were identified that had a 3-18-fold improvement in affinity (KD=15-88 nM) for EGFR-expressing A431 tumor cells compared to C10 scFv (KD=264 nM). By combining mutations, higher affinity scFv were generated with KD ranging from 0.9 nM to 10 nM. The highest affinity scFv had a 280-fold higher affinity compared to that of the parental C10 scFv. Immunoliposome nanoparticles (ILs) were prepared using EGFR scFv with a 280-fold range of affinities, and their binding and uptake into EGFR-expressing tumor cells was quantified. At scFv densities greater than 148 scFv/IL, there was no effect of scFv affinity on IL binding and uptake into tumor cells, or on cytotoxicity. At lower scFv densities, there was less uptake and binding for ILs constructed from the very low affinity C10 scFv. The results show the importance of antibody fragment density on nanoparticle uptake, and suggest that engineering ultrahigh affinity scFv may be unnecessary for optimal nanoparticle targeting. 相似文献
67.
Gritsenko VA Ivanov IuB Zhurlov OS 《Zhurnal mikrobiologii, epidemiologii, i immunobiologii》2006,(4):16-19
Testing 54 strains of staphylococci (Staphylococcus aureus, S. epidermidis, S. haemolyticus, S. warneri, S. hominis, S. capitis) revealed that S. aureus in contrast to coagulase-negative staphylococci (CNS) is more resistant to bactoriocidal action of human thrombodefensins (resistance index: 60.3 vs 25.6%), less hydrophilicolipophilic balance-HLB: -0.42 vs -0.64) and less charged (x-potential: -32.4 vs -35.6 mV). In groups of staphylococci (S. aureus and CNS) correlation links of bacterial resistance to human thrombodefensins with their HLB and x-potential (r=-0.32...-0.36). By In vitro experiments, it was shown that 5 passages of staphylococci in meat-peptonic broth with human thrombodefensins (50 mkg protein/ml) lead to adaptation of bacteria followed by the formation of resistance to cationic peptides from thrombocytes, a decrease of hydrophobicity and x-potencial. The role of physico-chemical properties in providing thrombodefensin-resistance of staphylococci as a developmental factor of infectious-and-inflammatory process and persistence of bacteria was confirmed with Salmonella infection. 相似文献
68.
M. V. Ivanov A. Yu. Lein Yu. M. Miller S. K. Yusupov N. V. Pimenov B. Wehrli I. I. Rusanov A. Zehnder 《Microbiology》2000,69(4):449-459
The isotopic composition of particulate organic carbon (POC) from the Black Sea deep-water zone was studied during a Russian-Swiss
expedition in May 1998. POC from the upper part of the hydrogen sulfide zone (the C-layer) was found to be considerably enriched
with the12C isotope, as compared to the POC of the oxycline and anaerobic zone. In the C-layer waters, the concurrent presence of dissolved
oxygen and hydrogen sulfide and an increased rate of dark CO2 fixation were recorded, suggesting that the change in the POC isotopic composition occurs at the expense of newly formed
isotopically light organic matter of the biomass of autotrophic bacteria involved in the sulfur cycle. In the anaerobic waters
below the C-layer, the organic matter of the biomass of autotrophs is consumed by the community of heterotrophic microorganisms;
this results in weighting of the POC isotopic composition. Analysis of the data obtained and data available in the literature
allows an inference to be made about the considerable seasonable variability of the POC δ13C value, which depends on the ratio of terrigenic and planktonogenic components in the particulate organic matter. 相似文献
69.
Krysko DV Diez-Fraile A Criel G Svistunov AA Vandenabeele P D'Herde K 《Apoptosis : an international journal on programmed cell death》2008,13(9):1065-1087
The vertebrate ovary is an extremely dynamic organ in which excessive or defective follicles are rapidly and effectively eliminated
early in ontogeny and thereafter continuously throughout reproductive life. More than 99% of follicles disappear, primarily
due to apoptosis of granulosa cells, and only a minute fraction of the surviving follicles successfully complete the path
to ovulation. The balance between signals for cell death and survival determines the destiny of the follicles. An abnormally
high rate of cell death followed by atresia can negatively affect fertility and eventually lead irreversibly to premature
ovarian failure. In this review we provide a short overview of the role of programmed cell death in prenatal differentiation
of the primordial germ cells and in postnatal folliculogenesis. We also discuss the issue of neo-oogenesis. Next, we highlight
molecules involved in regulation of granulosa cell apoptosis. We further discuss the potential use of scores for apoptosis
in granulosa cells and characteristics of follicular fluid as prognostic markers for predicting the outcome of assisted reproduction.
Potential therapeutic strategies for combating premature ovarian failure are also addressed. 相似文献
70.
Sergey Ivanov Maria J. Harrison 《The Plant journal : for cell and molecular biology》2014,80(6):1151-1163
Medicago truncatula is widely used for analyses of arbuscular mycorrhizal (AM) symbiosis and nodulation. To complement the genetic and genomic resources that exist for this species, we generated fluorescent protein fusions that label the nucleus, endoplasmic reticulum, Golgi apparatus, trans‐Golgi network, plasma membrane, apoplast, late endosome/multivesicular bodies (MVB), transitory late endosome/ tonoplast, tonoplast, plastids, mitochondria, peroxisomes, autophagosomes, plasmodesmata, actin, microtubules, periarbuscular membrane (PAM) and periarbuscular apoplastic space (PAS) and expressed them from the constitutive AtUBQ10 promoter and the AM symbiosis‐specific MtBCP1 promoter. All marker constructs showed the expected expression patterns and sub‐cellular locations in M. truncatula root cells. As a demonstration of their utility, we used several markers to investigate AM symbiosis where root cells undergo major cellular alterations to accommodate their fungal endosymbiont. We demonstrate that changes in the position and size of the nuclei occur prior to hyphal entry into the cortical cells and do not require DELLA signaling. Changes in the cytoskeleton, tonoplast and plastids also occur in the colonized cells and in contrast to previous studies, we show that stromulated plastids are abundant in cells with developing and mature arbuscules, while lens‐shaped plastids occur in cells with degenerating arbuscules. Arbuscule development and secretion of the PAM creates a periarbuscular apoplastic compartment which has been assumed to be continuous with apoplast of the cell. However, fluorescent markers secreted to the periarbuscular apoplast challenge this assumption. This marker resource will facilitate cell biology studies of AM symbiosis, as well as other aspects of legume biology. 相似文献