共查询到20条相似文献,搜索用时 15 毫秒
1.
Shibasaki S Kuroda K Duc Nguyen H Mori T Zou W Ueda M 《Applied microbiology and biotechnology》2006,70(4):451-457
A novel protein molecular targeting system was created using a cytoplasmic face of a plasma membrane-targeting system in Saccharomyces cerevisiae. The technique involves a molecular display for the creation of a novel reaction site and interaction sites in the field
of biotechnology. In a model system, a fluorescent protein was targeted as a reporter to the cytoplasmic face of the plasma
membrane. The C-terminal transmembrane domain (CTM) of Ras2p and Snc2p was examined as the portions with anchoring ability
to the cytoplasmic face of the plasma membrane. We found that the CTM of Snc2p targeted the enhanced cyan fluorescent protein
(ECFP)–protein A fusion protein on the cytoplasmic face of the plasma membrane more strongly than that of Ras2p. To develop
it for use as a detection system for protein–protein interactions, the Fc fragment of IgG (Fc) was genetically fused with
the enhanced yellow fluorescent protein (EYFP) and expressed in the cytoplasm of the ECFP–protein A-anchored cell. A microscopic
analysis showed that fluorescence resonance energy transfer (FRET) between ECFP–protein A and EYFP–Fc occurred, and the change
in fluorescence was observed on the cytoplasmic face of the plasma membrane. The detection of protein–protein interactions
at the cytoplasmic face of a plasma membrane using FRET combined with a cytoplasmic face-targeting system for proteins provides
a novel method for examining the molecular interactions of cytoplasmic proteins, in addition to conventional methods, such
as the two-hybrid method in the nuclei.
S. Shibasaki and K. Kuroda equally contributed to this work 相似文献
2.
Contessi S Comelli M Cmet S Lippe G Mavelli I 《Journal of bioenergetics and biomembranes》2007,39(4):291-300
F0F1ATPsynthase is now known to be expressed as a plasma membrane receptor for several extracellular ligands. On hepatocytes,
ecto–F0F1ATPsynthase binds apoA–I and triggers HDL endocytosis concomitant with ATP hydrolysis. Considering that inhibitor protein
IF1 was shown to regulate the hydrolytic activity of ecto–F0F1ATPsynthase and to interact with calmodulin (CaM) in vitro, we investigated the subcellular distributions of IF1, calmodulin (CaM), OSCP and β subunits of F0F1ATPsynthase in HepG2 cells. Using immunofluorescence and Western blotting, we found that around 50% of total cellular IF1 is localized outside mitochondria, a relevant amount of which is associated to the plasma membrane where we also found Ca2+–CaM, OSCP and β. Confocal microscopy showed that IF1 colocalized with Ca2+–CaM on plasma membrane but not in mitochondria, suggesting that Ca2+–CaM may modulate the cell surface availability of IF1 and thus its ability to inhibit ATP hydrolysis by ecto–F0F1ATPsynthase. These observations support a hypothesis that the IF1–Ca2+–CaM complex, forming on plasma membrane, functions in the cellular regulation of HDL endocytosis by hepatocytes. 相似文献
3.
Summary. Plants have evolved many mechanisms to cope with adverse environmental stresses. Abscisic acid (ABA) accumulates significantly
in plant cells in response to drought conditions, and this is believed to be a major mechanism through which plants enhance
drought tolerance. In this study, we explore the possible mechanisms of osmotic stress perception by plant cells and the consequent
induction of ABA biosynthesis. Immunoblotting and immunofluorescence localization experiments, using a polyclonal antibody
against human integrin β1, revealed the presence of a protein in Zea mays roots that is similar to the integrin proteins of animals and mainly localized in the plasma membrane. Treatment with GRGDS,
a synthetic pentapeptide containing an RGD domain, which interacted specifically with the integrin protein and thus blocked
the cell wall–plasma membrane interaction, significantly inhibited osmotic stress-induced ABA biosynthesis in cells, and the
GRGDS analog which does not contain the RGD domain had no effect. Our results show that a strong interaction exists between
the cell wall and plasma membrane and that this interaction is largely mediated by integrin-like proteins. They also imply
that the cell wall and/or cell wall–plasma membrane interaction plays important roles in the perception of osmotic stress.
Accordingly, we conclude that the cell wall and/or cell wall–plasma membrane interaction mediated by the integrin-like protein
plays important roles in osmotic stress-induced ABA biosynthesis in Zea mays.
Correspondence: J. S. Liang, College of Bioscience and Biotechnology, Yangzhou University, Yangzhou 225009, People’s Republic
of China. 相似文献
4.
Tandon C De Lisle RC Boulatnikov I Naik PK 《Molecular and cellular biochemistry》2007,302(1-2):157-167
The C-terminal PDZ-binding motifs are required for polarized apical/basolateral localization of many membrane proteins. Ezrin–radixin–moesin
(ERM) proteins regulate the organization and function of specific cortical structures in polarized epithelial cells by connecting
filamentous (F)-actin to plasma membrane proteins through EBP50. Previous work showed that the membrane phosphoprotein apactin
(an 80-kDa type I membrane protein derived from pro-Muclin) is associated with the acinar cell apical actin cytoskeleton and
that this association is modulated by changes in the phosphorylation state of the apactin cytosolic tail. The carboxyl-terminal
amino acids of apactin (–STKL–COOH) are predicted to form a type I PDZ-binding domain, similar to that of CFTR (–DTRL–COOH).
Pairwise sequence comparison between NHERF/EBP50 and PDZK1/CAP70 PDZ domains reveals significant identity among the 83 amino-acid
residues (12–92) of EBP50 and CAP70 (241–323), which are involved in the interaction with the carboxyl-terminal peptides (STKL–COOH
and phosphomimetics) of apactin. Hence, the specificity and affinity of interactions are identical between them, which is
corroborated with the two hybrid results. Substitution of all the four-carboxyl-terminal amino acids in the wild type to Ala
reduces the interaction. Only the carbonyl oxygen and amide nitrogen of Ala are found to be involved in hydrogen bonding.
Further, truncation of the wild carboxyl-terminal peptide to RGQPP–COOH, showed very low affinity of interaction with PDZ1
domain. Only the atom Oε1 of Gln-2 hydrogen bonds with Nε2 of His72 of PDZ domain. Ser-3 amino acid in wild type apactin protein (STKL–COOH) is not involved in hydrogen bonding with
PDZ1 domain. However, substitution of Ser-3 to Asp-3 in PDTKL–COOH peptide increases the affinity of interaction of PDTKL–COOH
with PDZ1 domain. Thus, carboxyl-terminal Asp(D) -3, Thr(T) -2, Lys(K) -1 and Leu(L) 0 are involved in numerous interactions
with PDZ1 domains of NHERF/EBP50 and PDZK1/CAP70. 相似文献
5.
Seven transmembrane domain G-protein-coupled receptors constitute the largest family of proteins in mammals. Signal transduction
events mediated by such receptors are the primary means by which cells communicate with and respond to their external environment.
The major paradigm in this signal transduction process is that stimulation of the receptor leads to the recruitment and activation
of heterotrimeric GTP-binding proteins. These initial events, which are fundamental to all types of G-protein-coupled receptor
signaling, occur at the plasma membrane via protein–protein interactions. As a result, the dynamics of the activated receptor
on cell surfaces represents an important determinant in its encounter with G-proteins, and has significant impact on the overall
efficiency of the signal transduction process. We have monitored the cell surface dynamics of the serotonin1A receptor, an important member of the G-protein-coupled receptor superfamily, in relation to its interaction with G-proteins.
Fluorescence recovery after photobleaching experiments carried out with the receptor tagged to the enhanced yellow fluorescent
protein indicate that G-protein activation alters the diffusion properties of the receptor in a manner suggesting the activation
process leads to dissociation of G-proteins from the receptor. This result demonstrates that the cell surface dynamics of
the serotonin1A receptor is modulated in a G-protein-dependent manner. Importantly, this result could provide the basis for a sensitive and
powerful approach to assess receptor/G-protein interaction in an intact cellular environment. 相似文献
6.
Carlos J. Baier Cristina E. Gallegos Valeria Levi Francisco J. Barrantes 《European biophysics journal : EBJ》2010,39(2):213-227
Nicotinic acetylcholine receptor (AChR) function and distribution are quite sensitive to cholesterol (Chol) levels in the
plasma membrane (reviewed by Barrantes in J Neurochem 103 (suppl 1):72–80, 2007). Here we combined confocal fluorescence recovery after photobleaching (FRAP) and confocal fluorescence correlation spectroscopy
(FCS) to examine the mobility of the AChR and its dependence on Chol content at the cell surface of a mammalian cell line.
Plasma membrane AChR exhibited limited mobility and only ~55% of the fluorescence was recovered within 10 min after photobleaching.
Depletion of membrane Chol by methyl-β-cyclodextrin strongly affected the mobility of the AChR at the plasma membrane; the
fraction of mobile AChR fell from 55 to 20% in Chol-depleted cells, whereas Chol enrichment by methyl-β-cyclodextrin-Chol
treatment did not reduce receptor mobility at the cell surface. Actin depolymerization caused by latrunculin A partially restored
receptor mobility in Chol-depleted cells. In agreement with the FRAP data, scanning FCS experiments showed that the diffusion
coefficient of the AChR was about 30% lower upon Chol depletion. Taken together, these results suggest that membrane Chol
modulates AChR mobility at the plasma membrane through a Chol-dependent mechanism sensitive to cortical actin. 相似文献
7.
(1) The serotonin1A receptor is a G-protein coupled receptor involved in several cognitive, behavioral, and developmental functions. It binds
the neurotransmitter serotonin and signals across the membrane through its interactions with heterotrimeric G-proteins. (2)
Lipid–protein interactions in membranes play an important role in the assembly, stability, and function of membrane proteins.
The role of membrane environment in serotonin1A receptor function is beginning to be addressed by exploring the consequences of lipid manipulations on the ligand binding
and G-protein coupling of serotonin1A receptors, the ability to functionally solubilize the serotonin1A receptor, and the factors influencing the membrane organization of the serotonin1A receptor. (3) Recent developments involving the application of detergent-based and detergent-free approaches to understand
the membrane organization of the serotonin1A receptor under conditions of ligand activation and modulation of membrane lipid content, with an emphasis on membrane cholesterol,
are described. 相似文献
8.
Braun M 《Protoplasma》2002,219(3-4):150-159
Summary. The noninvasive infrared laser micromanipulation technique (optical tweezers, optical trapping) and centrifugation were used
to study susception and perception, the early events in the gravitropic pathway of tip-growing characean rhizoids and protonemata.
Reorientation of the growth direction in both cell types was only initiated when at least 2–3 statoliths settled on specific
areas of the plasma membrane. This statolith-sensitive plasma membrane area is confined to the statolith region (10–35 μm
behind the tip) in positively gravitropic rhizoids, whereas in negatively gravitropic protonemata, this area is limited to
the apical plasma membrane (0–10 μm). Statolith sedimentation towards the sensitive plasma membrane areas is mediated by the
concerted action of actin and gravity. The process of sedimentation, the pure physical movement, of statoliths is not sufficient
to initiate graviresponses in both cell types. It is concluded that specific statolith-sensitive plasma membrane areas play
a crucial role in the signal transduction pathway of gravitropism. These areas may represent the primary sites for gravity
perception and may transform the information derived from the gravity-induced statolith sedimentation into physiological signals
which trigger the molecular mechanisms of the opposite graviresponses in characean rhizoids and protonemata.
Received September 10, 2001 Accepted November 16, 2001 相似文献
9.
Karl-Ernst Kaissling 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》2009,195(10):895-922
Modelling reveals that within about 3 ms after entering the sensillum lymph, 17% of total pheromone is enzymatically degraded
while 83% is bound to the pheromone-binding protein (PBP) and thereby largely protected from enzymatic degradation. The latter
proceeds within minutes, 20,000-fold more slowly than with the free pheromone. In vivo the complex pheromone–PBP interacts
with the receptor molecule. At weak stimulation the half-life of the active complex is 0.8 s due to the postulated pheromone
deactivation. Most likely this process is enzymatically catalysed; it changes the PBP into a scavenger form, possibly by interference
with the C-terminus. The indirectly determined PBP concentration (3.8 mM) is close to direct measurements. The calculated
density of receptor molecules within the plasma membrane of the receptor neuron reaches up to 6,000 units per μm2. This is compared with the estimated densities of the sensory-neuron membrane protein and of ion channels. The EC50 of the model pheromone–PBP complex interacting with the receptor molecules is 6.8 μM, as compared with the EC50 = 1.5 μM of bombykol recently determined using heterologous expression. A possible mechanism widening the range of stimulus
intensities covered by the dose–response curve of the receptor-potential is proposed. 相似文献
10.
A hypothesis for the minimal overall structure of the mammalian plasma membrane redox system 总被引:2,自引:0,他引:2
de Grey AD 《Protoplasma》2003,221(1-2):3-9
Summary. After a long period of frustration, many components of the mammalian plasma membrane redox system are now being identified
at the molecular level. Some are apparently ubiquitous but are necessary only for a subset of electron donors or acceptors;
some are present only in certain cell types; some appear to be associated with proton extrusion; some appear to be capable
of superoxide production. The volume and variety of data now available have begun to allow the formulation of tentative models
for the overall network of interactions of enzymes and substrates that together make up the plasma membrane redox system.
Such a model is presented here. The structure discussed here is of the mammalian system, though parts of it may apply more
or less accurately to fungal and plant cells too. Judging from the history of mitochondrial oxidative phosphorylation, it
may be hoped that the development of models of the whole system – even if they undergo substantial revision thereafter – will
markedly accelerate the pace of research in plasma membrane redox, by providing a coherent basis for the design of future
experiments.
Received May 4, 2002; accepted July 26, 2002; published online May 21, 2003
RID="*"
ID="*" Correspondence and reprints: Department of Genetics, University of Cambridge, Downing Street, Cambridge CB2 3EH, United
Kingdom. E-mail: ag24@gen.cam.ac.uk 相似文献
11.
The serotonin1A receptor is the most extensively studied member of the family of seven transmembrane domain G-protein coupled serotonin receptors. Since a large portion of such transmembrane receptors remains in contact with the membrane lipid environment, lipid–protein interactions assume importance in the structure-function analysis of such receptors. We have earlier reported the requirement of cholesterol for serotonin1A receptor function in native hippocampal membranes by specific depletion of cholesterol using methyl- β-cyclodextrin. In this paper, we monitored the serotonin1A receptor function in membranes that are enriched in cholesterol using a complex prepared from cholesterol and methyl-β-cyclodextrin. Our results indicate that ligand binding and receptor/G-protein interaction of the serotonin1A receptor do not exhibit significant difference in native and cholesterol-enriched hippocampal membranes indicating that further enrichment of cholesterol has little functional consequence on the serotonin1A receptor function. These results therefore provide new information on the effect of cholesterol enrichment on the hippocampal serotonin1A receptor function. 相似文献
12.
The physiological relationship between brain cholesterol content and the action of amyloid β (Aβ) peptide in Alzheimer’s disease (AD) is a highly controversially discussed topic. Evidences for modulations of the Aβ/membrane interaction induced by plasma membrane cholesterol have already been observed. We have recently reported that Aβ(25–35) is capable of inserting in lipid membranes and perturbing their structure. Applying neutron diffraction and selective deuteration, we now demonstrate that cholesterol alters, at the molecular level, the capability of Aβ(25–35) to penetrate into the lipid bilayers; in particular, a molar weight content of 20% of cholesterol hinders the intercalation of monomeric Aβ(25–35) completely. At very low cholesterol content (about 1% molar weight) the location of the C-terminal part of Aβ(25–35) has been unequivocally established in the hydrocarbon region of the membrane, in agreement with our previous results on pure phospholipids membrane. These results link a structural property to a physiological and functional behavior and point to a therapeutical approach to prevent the AD by modulation of membrane properties. 相似文献
13.
P2X receptor (P2XR) is a family of the ATP-gated ion channel family and can permeabilize the plasma membrane to small cations
such as potassium, sodium, and calcium, resulting in cellular depolarization. There are seven P2XR that have been described
and cloned, with 45% identity in amino acid sequence. Each P2X receptors has two transmembrane domains that are separated
by an extracellular loop and an intracellular N and C terminus. Unlike the other P2X receptors, the P2X7R has a larger C terminus
with an extra 200 amino acid residues compared with the other receptors. The C terminus of the P2X7R has been implicated in
regulating receptor function including signaling pathway activation, cellular localization, protein–protein interactions,
and post-translational modification (PTM). In the present review, we discuss the role of the P2X7R C terminus in regards to
receptor function, describe the specific domains and motifs found therein and compare the C terminus sequence with others
proteins to discover predicted domains or sites of PTM. 相似文献
14.
The platelet receptor for type III collagen (TIIICBP) is present in platelet membrane lipid microdomains (rafts) 总被引:2,自引:2,他引:0
Maurice P Waeckel L Pires V Sonnet P Lemesle M Arbeille B Vassy J Rochette J Legrand C Fauvel-Lafève F 《Histochemistry and cell biology》2006,125(4):407-417
Platelet interactions with collagen are orchestrated by the presence or the migration of platelet receptor(s) for collagen
into lipid rafts, which are specialized lipid microdomains from the platelet plasma membrane enriched in signalling proteins.
Electron microscopy shows that in resting platelets, TIIICBP, a receptor specific for type III collagen, is present on the
platelet membrane and associated with the open canalicular system, and redistributes to the platelet membrane upon platelet
activation. After platelet lysis by 1% Triton X-100 and the separation of lipid rafts on a discontinuous sucrose gradient,
TIIICBP is recovered in lipid raft-containing fractions and Triton X-100 insoluble fractions enriched in cytoskeleton proteins.
Platelet aggregation, induced by type III collagen, was inhibited after disruption of the lipid rafts by cholesterol depletion,
whereas platelet adhesion under static conditions did not require lipid raft integrity. These results indicate that TIIICBP,
a platelet receptor involved in platelet interaction with type III collagen, is localized within platelet lipid rafts where
it could interact with other platelet receptors for collagen (GP VI and α2β1 integrin) for efficient platelet activation.
Pascal Maurice and Ludovic Waeckel have contributed equally to this work. 相似文献
15.
Mannose 6-phosphate receptor proteins (MPR 300 and 46) in mammals have been shown to mediate transport of lysosomal enzymes to lysosomes intracellularly. Both receptors are also expressed on the plasma membrane. Only MPR 300 protein on the plasma membrane has been shown to be a multifunctional protein which in addition to binding mannose 6-phosphate containing proteins also binds human insulin-like growth factor-II (IGF-II) causing its internalization [Hille-Rehfeld, A. (1995) Mannose 6-phosphate receptors in sorting and transport of lysosomal enzymes. Biochim. Biophys. Acta. 1241: 177–194]. This property has been shown to be exhibited by other mammalian receptors but not by the chicken and frog receptors. In a recent study however it was shown that the fish embryo MPR 300 binds human IGF-II. [Mendez, E., Planas, J.V., Castillo, J., Navarro, I. and Gutierrez, J. (2001) Identification of a type II insulin-like growth factor receptor in fish embryos. Endocrinology, 142: 1090–1097]. In the present study, we demonstrate that the purified goat and chicken liver receptors bind human IGF-II by employing cross-linking experiments (purified receptors and radiolabeled IGF-II) and by ligand blotting (using purified receptors and biotinylated IGF-II). Further CEF cells (chicken embryonic fibroblasts) that are known to contain the putative MPR 300 protein were employed to demonstrate that the CEF cell receptor binds human IGF-II. 相似文献
16.
Cholesterol is an essential component of eukaryotic membranes and plays a crucial role in membrane organization, dynamics
and function. The G-protein coupled receptors (GPCRs) are the largest class of molecules involved in signal transduction across
membranes and constitute ~1–2% of the human genome. GPCRs have emerged as major targets for the development of novel drug
candidates in all clinical areas due to their involvement in the generation of multitude of cellular responses. Membrane cholesterol
has been reported to have a modulatory role in the function of a number of GPCRs. This effect could either be due to specific
molecular interaction between cholesterol and GPCR, or due to alterations in the membrane physical properties induced by cholesterol.
Alternatively, membrane cholesterol could modulate receptor function by occupying the ‘nonannular’ sites around the receptor.
In this review, we have highlighted the nature of cholesterol dependence of GPCR function taking a few known examples. 相似文献
17.
The analysis of protein–protein interactions is important for developing a better understanding of the functional annotations of proteins that are involved in various biochemical reactions in vivo. The discovery that a protein with an unknown function binds to a protein with a known function could provide a significant clue to the cellular pathway concerning the unknown protein. Therefore, information on protein–protein interactions obtained by the comprehensive analysis of all gene products is available for the construction of interactive networks consisting of individual protein–protein interactions, which, in turn, permit elaborate biological phenomena to be understood. Systems for detecting protein–protein interactions in vitro and in vivo have been developed, and have been modified to compensate for limitations. Using these novel approaches, comprehensive and reliable information on protein–protein interactions can be determined. Systems that permit this to be achieved are described in this review.K. Kuroda, M. Kato and J. Mima contributed equally to this work. 相似文献
18.
Cells in living organisms are regulated by chemical and physical stimuli from their environment. Often, ligands interact with
membrane receptors to trigger responses and Sargent and Schwyzer conceived a model to describe this process, “membrane catalysis”.
There is a notion that the physical organization of membranes can control the response of cells by speeding up reactions.
We revisit the “membrane catalysis” model in the light of recent technical, methodological and theoretical advances and how
they can be exploited to highlight the details of membrane mediated ligand–receptor interactions. We examine the possible
effects that ligand concentration causes in the membrane catalysis and focus our attention in techniques used to determine
the partition constant. The hypothetical diffusional advantage associated with membrane catalysis is discussed and the applicability
of existing models is assessed. The role of in-depth location and orientation of ligands is explored emphasizing the contribution
of new analysis methods and spectroscopic techniques. Results suggest that membranes can optimize the interaction between
ligands and receptors through several different effects but the relative contribution of each must be carefully investigated.
We certainly hope that the conjugation of the methodological and technical advances here reported will revive the interest
in the membrane catalysis model. 相似文献
19.
Lucie Khemtémourian Maarten F. M. Engel John A. W. Kruijtzer Jo W. M. Höppener Rob M. J. Liskamp J. Antoinette Killian 《European biophysics journal : EBJ》2010,39(9):1359-1364
Human islet amyloid polypeptide (hIAPP) forms amyloid fibrils in pancreatic islets of patients with type 2 diabetes mellitus.
It has been suggested that the N-terminal part, which contains a conserved intramolecular disulfide bond between residues
2 and 7, interacts with membranes, ultimately leading to membrane damage and β-cell death. Here, we used variants of the hIAPP1–19 fragment and model membranes of phosphatidylcholine and phosphatidylserine (7:3, molar ratio) to examine the role of this
disulfide in membrane interactions. We found that the disulfide bond has a minor effect on membrane insertion properties and
peptide conformational behavior, as studied by monolayer techniques, 2H NMR, ThT-fluorescence, membrane leakage, and CD spectroscopy. The results suggest that the disulfide bond does not play
a significant role in hIAPP–membrane interactions. Hence, the fact that this bond is conserved is most likely related exclusively
to the biological activity of IAPP as a hormone. 相似文献
20.
Jasionowski Marek Łankiewicz Leszek Wiczk Wiesław Gwizdała Elwira Grzonka Zbigniew 《International journal of peptide research and therapeutics》1998,5(5-6):371-374
Summary Conformational features of the neuropeptide pituitary adenylate cyclase activating polypeptide (1–27) (PACAP(1–27)) and its
shorter fragments (1–5), (7–11) and (14–27) were studied by circular dichroism (CD) and fluorescence spectroscopy. The obtained
CD spectra revealed that only PACAP(1–27) and the fragment (14–27) possess some content of an organized structure — the α-helix.
This C-terminal, helical part of the peptides is important for receptor binding as it provides a stable structure that can
reside in the ordered lipid region of the receptor site in the membrane, while the primary biological function of the hormone
resides in the N-terminal, disordered part. Fluorescence studies have revealed that the tyrosine residue located in the helical
region of PACAP has a higher quantum yield and a longer average lifetime than the tyrosine in the N-terminus, probably due
to a ‘shielding’ effect of the hydrophobic cluster around Tyr22. 相似文献