共查询到20条相似文献,搜索用时 31 毫秒
1.
Qi Guo Lei Liu Won C. Yim John C. Cushman Bronwyn J. Barkla 《International journal of molecular sciences》2021,22(9)
The study of subcellular membrane structure and function facilitates investigations into how biological processes are divided within the cell. However, work in this area has been hampered by the limited techniques available to fractionate the different membranes. Free Flow Electrophoresis (FFE) allows for the fractionation of membranes based on their different surface charges, a property made up primarily of their varied lipid and protein compositions. In this study, high-resolution plant membrane fractionation by FFE, combined with mass spectrometry-based proteomics, allowed the simultaneous profiling of multiple cellular membranes from the leaf tissue of the plant Mesembryanthemum crystallinum. Comparisons of the fractionated membranes’ protein profile to that of known markers for specific cellular compartments sheds light on the functions of proteins, as well as provides new evidence for multiple subcellular localization of several proteins, including those involved in lipid metabolism. 相似文献
2.
Minu Chaudhuri Anuj Tripathi Fidel Soto Gonzalez 《International journal of molecular sciences》2021,22(15)
Mitochondria are essential in eukaryotes. Besides producing 80% of total cellular ATP, mitochondria are involved in various cellular functions such as apoptosis, inflammation, innate immunity, stress tolerance, and Ca2+ homeostasis. Mitochondria are also the site for many critical metabolic pathways and are integrated into the signaling network to maintain cellular homeostasis under stress. Mitochondria require hundreds of proteins to perform all these functions. Since the mitochondrial genome only encodes a handful of proteins, most mitochondrial proteins are imported from the cytosol via receptor/translocase complexes on the mitochondrial outer and inner membranes known as TOMs and TIMs. Many of the subunits of these protein complexes are essential for cell survival in model yeast and other unicellular eukaryotes. Defects in the mitochondrial import machineries are also associated with various metabolic, developmental, and neurodegenerative disorders in multicellular organisms. In addition to their canonical functions, these protein translocases also help maintain mitochondrial structure and dynamics, lipid metabolism, and stress response. This review focuses on the role of Tim50, the receptor component of one of the TIM complexes, in different cellular functions, with an emphasis on the Tim50 homologue in parasitic protozoan Trypanosoma brucei. 相似文献
3.
Membrane proteins evolved to reside in the hydrophobic lipid bilayers of cellular membranes. Therefore, membrane proteins bridge the different aqueous compartments separated by the membrane, and furthermore, dynamically interact with their surrounding lipid environment. The latter not only stabilizes membrane proteins, but directly impacts their folding, structure and function. In order to be characterized with biophysical and structural biological methods, membrane proteins are typically extracted and subsequently purified from their native lipid environment. This approach requires that lipid membranes are replaced by suitable surrogates, which ideally closely mimic the native bilayer, in order to maintain the membrane proteins structural and functional integrity. In this review, we survey the currently available membrane mimetic environments ranging from detergent micelles to bicelles, nanodiscs, lipidic-cubic phase (LCP), liposomes, and polymersomes. We discuss their respective advantages and disadvantages as well as their suitability for downstream biophysical and structural characterization. Finally, we take a look at ongoing methodological developments, which aim for direct in-situ characterization of membrane proteins within native membranes instead of relying on membrane mimetics. 相似文献
4.
Aritz B. García-Arribas Flix M. Goi Alicia Alonso 《International journal of molecular sciences》2021,22(18)
Lipid model membranes are important tools in the study of biophysical processes such as lipid self-assembly and lipid–lipid interactions in cell membranes. The use of model systems to adequate and modulate complexity helps in the understanding of many events that occur in cellular membranes, that exhibit a wide variety of components, including lipids of different subfamilies (e.g., phospholipids, sphingolipids, sterols…), in addition to proteins and sugars. The capacity of lipids to segregate by themselves into different phases at the nanoscale (nanodomains) is an intriguing feature that is yet to be fully characterized in vivo due to the proposed transient nature of these domains in living systems. Model lipid membranes, instead, have the advantage of (usually) greater phase stability, together with the possibility of fully controlling the system lipid composition. Atomic force microscopy (AFM) is a powerful tool to detect the presence of meso- and nanodomains in a lipid membrane. It also allows the direct quantification of nanomechanical resistance in each phase present. In this review, we explore the main kinds of lipid assemblies used as model membranes and describe AFM experiments on model membranes. In addition, we discuss how these assemblies have extended our knowledge of membrane biophysics over the last two decades, particularly in issues related to the variability of different model membranes and the impact of supports/cytoskeleton on lipid behavior, such as segregated domain size or bilayer leaflet uncoupling. 相似文献
5.
Naghmeh Poorinmohammad Jing Fu Bob Wabeke Eduard J. Kerkhoven 《International journal of molecular sciences》2022,23(15)
Given the strong potential of Yarrowia lipolytica to produce lipids for use as renewable fuels and oleochemicals, it is important to gain in-depth understanding of the molecular mechanism underlying its lipid accumulation. As cellular growth rate affects biomass lipid content, we performed a comparative proteomic analysis of Y. lipolytica grown in nitrogen-limited chemostat cultures at different dilution rates. After confirming the correlation between growth rate and lipid accumulation, we were able to identify various cellular functions and biological mechanisms involved in oleaginousness. Inspection of significantly up- and downregulated proteins revealed nonintuitive processes associated with lipid accumulation in this yeast. This included proteins related to endoplasmic reticulum (ER) stress, ER–plasma membrane tether proteins, and arginase. Genetic engineering of selected targets validated that some genes indeed affected lipid accumulation. They were able to increase lipid content and were complementary to other genetic engineering strategies to optimize lipid yield. 相似文献
6.
António M. T. Martins do Canto Alfredo J. Palace Carvalho Jo?o P. Prates Ramalho Luís M. S. Loura 《International journal of molecular sciences》2013,14(7):14724-14743
T-20 and T-1249 fusion inhibitor peptides were shown to interact with 1-palmitoyl-2-oleyl-phosphatidylcholine (POPC) (liquid disordered, ld) and POPC/cholesterol (1:1) (POPC/Chol) (liquid ordered, lo) bilayers, and they do so to different extents. Although they both possess a tryptophan-rich domain (TRD), T-20 lacks a pocket binding domain (PBD), which is present in T-1249. It has been postulated that the PBD domain enhances FI interaction with HIV gp41 protein and with model membranes. Interaction of these fusion inhibitor peptides with both the cell membrane and the viral envelope membrane is important for function, i.e., inhibition of the fusion process. We address this problem with a molecular dynamics approach focusing on lipid properties, trying to ascertain the consequences and the differences in the interaction of T-20 and T-1249 with ld and lo model membranes. T-20 and T-1249 interactions with model membranes are shown to have measurable and different effects on bilayer structural and dynamical parameters. T-1249’s adsorption to the membrane surface has generally a stronger influence in the measured parameters. The presence of both binding domains in T-1249 appears to be paramount to its stronger interaction, and is shown to have a definite importance in membrane properties upon peptide adsorption. 相似文献
7.
Dr. Gakushi Tsuji Dr. Takeshi Sunami Dr. Masaya Oki Dr. Norikazu Ichihashi 《Chembiochem : a European journal of chemical biology》2021,22(11):1966-1973
Liposomes, which are vesicles surrounded by lipid membranes, can be used as biochemical reactors by encapsulating various reactions. Accordingly, they are useful for studying cellular functions under controlled conditions that mimic the environment within a cell. However, one of the shortcomings of liposomes as biochemical reactors is the difficulty of introducing or removing proteins due to the impermeability of the membrane. In this study, we established a method for exchanging proteins in liposomes by forming reversible pores in the membrane. We used the toxic protein streptolysin O (SLO); this forms pores in membranes made of phospholipids containing cholesterol that can be closed by the addition of calcium ions. After optimizing the experimental procedure and lipid composition, we observed the exchange of fluorescent proteins (transferrin Alexa Fluor 488 and 647) in 9.9 % of liposomes. We also introduced T7 RNA polymerase, a 98-kDa enzyme, and observed RNA synthesis in ∼8 % of liposomes. Our findings establish a new method for controlling the internal protein composition of liposomes, thereby increasing their utility as bioreactors. 相似文献
8.
Bipolar tetraether lipids (BTL) have been long thought to play a critical role in allowing thermoacidophiles to thrive under extreme conditions. In the present study, we demonstrated that not all BTLs from the thermoacidophilic archaeon Sulfolobus acidocaldarius exhibit the same membrane behaviors. We found that free-standing planar membranes (i.e., black lipid membranes, BLM) made of the polar lipid fraction E (PLFE) isolated from S. acidocaldarius formed over a pinhole on a cellulose acetate partition in a dual-chamber Teflon device exhibited remarkable stability showing a virtually constant capacitance (~28 pF) for at least 11 days. PLFE contains exclusively tetraethers. The dominating hydrophobic core of PLFE lipids is glycerol dialky calditol tetraether (GDNT, ~90%), whereas glycerol dialkyl glycerol tetraether (GDGT) is a minor component (~10%). In sharp contrast, BLM made of BTL extracted from microvesicles (Sa-MVs) released from the same cells exhibited a capacitance between 36 and 39 pF lasting for only 8 h before membrane dielectric breakdown. Lipids in Sa-MVs are also exclusively tetraethers; however, the dominating lipid species in Sa-MVs is GDGT (>99%), not GDNT. The remarkable stability of BLMPLFE can be attributed to strong PLFE–PLFE and PLFE–substrate interactions. In addition, we compare voltage-dependent channel activity of calcium-gated potassium channels (MthK) in BLMPLFE to values recorded in BLMSa-MV. MthK is an ion channel isolated from a methanogenic that has been extensively characterized in diester lipid membranes and has been used as a model for calcium-gated potassium channels. We found that MthK can insert into BLMPLFE and exhibit channel activity, but not in BLMSa-MV. Additionally, the opening/closing of the MthK in BLMPLFE is detectable at calcium concentrations as low as 0.1 mM; conversely, in diester lipid membranes at such a low calcium concentration, no MthK channel activity is detectable. The differential effect of membrane stability and MthK channel activity between BLMPLFE and BLMSa-MV may be attributed to their lipid structural differences and thus their abilities to interact with the substrate and membrane protein. Since Sa-MVs that bud off from the plasma membrane are exclusively tetraether lipids but do not contain the main tetraether lipid component GDNT of the plasma membrane, domain segregation must occur in S. acidocaldarius. The implication of this study is that lipid domain formation is existent and functionally essential in all kinds of cells, but domain formation may be even more prevalent and pronounced in hyperthermophiles, as strong domain formation with distinct membrane behaviors is necessary to counteract randomization due to high growth temperatures while BTL in general make archaea cell membranes stable in high temperature and low pH environments whereas different BTL domains play different functional roles. 相似文献
9.
A new concept of cellular uptake and intracellular trafficking of long-chain fatty acids 总被引:6,自引:0,他引:6
Fatty acids are the main structural and energy sources of the human body. Within the organism, they are presented to cells
as fatty acid: albumin complexes. Dissociation from albumin represents the first step of the cellular uptake process, involving
membrane proteins with high affinity for fatty acids, e.g., fatty acid translocase (FAT/CD 36) or the membrane fatty acid-binding
protein (FABPpm). According to the thus created transmembrane concentration gradient, uncharged fatty acids can flip-flop from the outer
leaflet across the phospholipid bilayer. At the cytosolic surface of the plasma membrane, fatty acids can associate with the
cytosolic FABP (FABP
c
) or with caveolin-1. Caveolins are constituents of caveolae, which are proposed to serve as lipid delivery vehicles for subcellular
organelles. It is not known whether protein (FABP
c
)- and lipid (caveolae)-mediated intracellular trafficking of fatty acids operates in conjunction, or in parallel. Channeling
fatty acids to the different metabolic pathways requires activation to acyl-CoA. For this process, the family of fatty acid
transport proteins (FATP 1-5/6) might be relevant because they have been shown to possess acyl-CoA synthetase activity. Their
variable N-terminal signaling sequences suggest that they might be targeted to specific organelles by anchoring in the phospholipid
bilayer of the different subcellular membranes. At the highly conserved cytosolic AMP-binding site of FATP, fatty acids are
activated to acyl-CoA for subsequent metabolic disposition by specific organelles. Overall, fatty acid uptake represents a
continuous flow involving the following: dissociation from albumin by membrane proteins with high affinity for fatty acids;
passive flip-flop across the phospholipid bilayer; binding to FABP
c
and caveolin-1 at the cytosolic plasma membrane; and intracellular trafficking via FABP
c
and/or caveolae to sites of metabolic disposition. The uptake process is terminated after activation to acyl-CoA by the members
of the FATP family targeted intracellularly to different organelles. 相似文献
10.
Dr. Abhishek Cukkemane Prof. Dr. Marc Baldus 《Chembiochem : a European journal of chemical biology》2013,14(14):1789-1798
Voltage‐gated ion channels are large tetrameric multidomain membrane proteins that play crucial roles in various cellular transduction pathways. Because of their large size and domain‐related mobility, structural characterization has proved challenging. We analyzed high‐resolution solid‐state NMR data on different isotope‐labeled protein constructs of a bacterial cyclic nucleotide‐activated K+ channel (MlCNG) in lipid bilayers. We could identify the different subdomains of the 4×355 residue protein, such as the voltage‐sensing domain and the cyclic nucleotide binding domain. Comparison to ssNMR data obtained on isotope‐labeled cell membranes suggests a tight association of negatively charged lipids to the channel. We detected spectroscopic polymorphism that extends beyond the ligand binding site, and the corresponding protein segments have been associated with mutant channel types in eukaryotic systems. These findings illustrate the potential of ssNMR for structural investigations on large membrane‐embedded proteins, even in the presence of local disorder. 相似文献
11.
Lo KM; Sudo Y; Chen J; Li Y; Lan Y; Kong SM; Chen L; An Q; Gillies SD 《Protein engineering, design & selection : PEDS》1998,11(6):495-500
We have developed a general expression system that enhances the production
and secretion of proteins in mammalian cells. The protein of interest is
expressed as a fusion to a signal peptide and the Fc fragment of
immunoglobulin as the N-terminal fusion partner, which can direct the
cellular processes into expressing and secreting high levels of many
different types of proteins. These include secretory proteins, enzymes and
soluble domains of membrane proteins, as well as nuclear and regulatory
proteins. Typical expression levels of these proteins from stable cell
lines ranged from several to 100 microg/ml in conditioned media. The Fc
domain helps to solubilize hydrophobic proteins and provides a handle for
easy detection and purification of the fusion proteins; and it can be
cleaved off by treatment with protease if desired.
相似文献
12.
Samantha Katarzyna Dziurdzik Elizabeth Conibear 《International journal of molecular sciences》2021,22(6)
The conserved VPS13 proteins constitute a new family of lipid transporters at membrane contact sites. These large proteins are suspected to bridge membranes and form a direct channel for lipid transport between organelles. Mutations in the 4 human homologs (VPS13A–D) are associated with a number of neurological disorders, but little is known about their precise functions or the relevant contact sites affected in disease. In contrast, yeast has a single Vps13 protein which is recruited to multiple organelles and contact sites. The yeast model system has proved useful for studying the function of Vps13 at different organelles and identifying the localization determinants responsible for its membrane targeting. In this review we describe recent advances in our understanding of VPS13 proteins with a focus on yeast research. 相似文献
13.
Protein lipidation and lipid trafficking control many key biological functions in all kingdoms of life. The discovery of diverse lipid species and their covalent attachment to many proteins has revealed a complex and regulated network of membranes and lipidated proteins that are central to fundamental aspects of physiology and human disease. Given the complexity of lipid trafficking and the protein targeting mechanisms involved with membrane lipids, precise and sensitive methods are needed to monitor and identify these hydrophobic molecules in bacteria, yeast, and higher eukaryotes. Although many analytical methods have been developed for characterizing membrane lipids and covalently modified proteins, traditional reagents and approaches have limited sensitivity, do not faithfully report on the lipids of interest, or are not readily accessible. The invention of bioorthogonal ligation reactions, such as the Staudinger ligation and azide-alkyne cycloadditions, has provided new tools to address these limitations, and their use has begun to yield fresh insight into the biology of protein lipidation and lipid trafficking. In this Account, we discuss how these new bioorthogonal ligation reactions and lipid chemical reporters afford new opportunities for exploring the biology of lipid-modified proteins and lipid trafficking. Lipid chemical reporters from our laboratory and several other research groups have enabled improved detection and large-scale proteomic analysis of fatty-acylated and prenylated proteins. For example, fatty acid and isoprenoid chemical reporters in conjunction with bioorthogonal ligation methods have circumvented the limited sensitivity and hazards of radioactive analogues, allowing rapid and robust fluorescent detection of lipidated proteins in all organisms tested. These chemical tools have revealed alterations in protein lipidation in different cellular states and are beginning to provide unique insights in mechanisms of regulation. Notably, the purification of proteins labeled with lipid chemical reporters has allowed both the large-scale analysis of lipidated proteins as well as the discovery of new lipidated proteins involved in metabolism, gene expression, and innate immunity. Specific lipid reporters have also been developed to monitor the trafficking of soluble lipids; these species are enabling bioorthogonal imaging of membranes in cells and tissues. Future advances in bioorthogonal chemistry, specific lipid reporters, and spectroscopy should provide important new insight into the functional roles of lipidated proteins and membranes in biology. 相似文献
14.
Hsin-Hui Shen Trevor Lithgow Lisandra L. Martin 《International journal of molecular sciences》2013,14(1):1589-1607
The function of any given biological membrane is determined largely by the specific set of integral membrane proteins embedded in it, and the peripheral membrane proteins attached to the membrane surface. The activity of these proteins, in turn, can be modulated by the phospholipid composition of the membrane. The reconstitution of membrane proteins into a model membrane allows investigation of individual features and activities of a given cell membrane component. However, the activity of membrane proteins is often difficult to sustain following reconstitution, since the composition of the model phospholipid bilayer differs from that of the native cell membrane. This review will discuss the reconstitution of membrane protein activities in four different types of model membrane—monolayers, supported lipid bilayers, liposomes and nanodiscs, comparing their advantages in membrane protein reconstitution. Variation in the surrounding model environments for these four different types of membrane layer can affect the three-dimensional structure of reconstituted proteins and may possibly lead to loss of the proteins activity. We also discuss examples where the same membrane proteins have been successfully reconstituted into two or more model membrane systems with comparison of the observed activity in each system. Understanding of the behavioral changes for proteins in model membrane systems after membrane reconstitution is often a prerequisite to protein research. It is essential to find better solutions for retaining membrane protein activities for measurement and characterization in vitro. 相似文献
15.
Berta Gorospe Jos J. G. Moura Carlos Gutierrez-Merino Alejandro K. Samhan-Arias 《International journal of molecular sciences》2022,23(23)
Caveolin-2 is a protein suitable for the study of interactions of caveolins with other proteins and lipids present in caveolar lipid rafts. Caveolin-2 has a lower tendency to associate with high molecular weight oligomers than caveolin-1, facilitating the study of its structural modulation upon association with other proteins or lipids. In this paper, we have successfully expressed and purified recombinant human caveolin-2 using E. coli. The structural changes of caveolin-2 upon interaction with a lipid bilayer of liposomes were characterized using bioinformatic prediction models, circular dichroism, differential scanning calorimetry, and fluorescence techniques. Our data support that caveolin-2 binds and alters cholesterol-rich domains in the membranes through a CARC domain, a type of cholesterol-interacting domain in its sequence. The far UV-CD spectra support that the purified protein keeps its folding properties but undergoes a change in its secondary structure in the presence of lipids that correlates with the acquisition of a more stable conformation, as shown by differential scanning calorimetry experiments. Fluorescence experiments using egg yolk lecithin large unilamellar vesicles loaded with 1,6-diphenylhexatriene confirmed that caveolin-2 adsorbs to the membrane but only penetrates the core of the phospholipid bilayer if vesicles are supplemented with 30% of cholesterol. Our study sheds light on the caveolin-2 interaction with lipids. In addition, we propose that purified recombinant caveolin-2 can provide a new tool to study protein–lipid interactions within caveolae. 相似文献
16.
Simon J. Slater Mary Beth Kelly Mark D. Yeager J. Larkin Cojen Ho Christopher D. Stubbs 《Lipids》1996,31(1):S189-S192
The effect of variation of the degree ofcis-unsaturation on cell membrane protein functioning was investigated using a model lipid bilayer system and protein kinase
C (PKC). This protein is a key element of signal transduction. Furthermore it is representative of a class of extrinsic membrane
proteins that show lipid dependent interactions with cell membranes. To test for dependence of activity on the phospholipid
unsaturation, experiments were devised using a vesicle assay system consisting of phosphatidylcholine (PC) and phosphatidylserine
(PS) in which the unsaturation was systematically varied. Highly purified PKCα and ε were obtained using the baculovirus-insect
cell expression system. It was shown that increased PC unsaturation elevated the activity of PKCα. By contrast, increasing
the unsaturation of PSdecreased the activity of PKCα, and to a lesser extent PKCε. This result immediately rules out any single lipid bilayer physical parameter,
such as lipid order, underlying the effect. It is proposed that while PC unsaturation effects are explainable on the basis
of a contribution to membrane surface curvature stress, the effects of PS unsaturation may be due to specific protein-lipid
interactions. Overall, the results indicate that altered phospholipid unsaturation in cell membranes that occurs in certain
disease states such as chronic alcoholism, or by dietary manipulations, are likely to have profound effects on signal transduction
pathways involving PKC and similar proteins. 相似文献
17.
Muhammad Shuja Khan Noura Sayed Dosoky John Dalton Williams 《International journal of molecular sciences》2013,14(11):21561-21597
Lipid membranes regulate the flow of nutrients and communication signaling between cells and protect the sub-cellular structures. Recent attempts to fabricate artificial systems using nanostructures that mimic the physiological properties of natural lipid bilayer membranes (LBM) fused with transmembrane proteins have helped demonstrate the importance of temperature, pH, ionic strength, adsorption behavior, conformational reorientation and surface density in cellular membranes which all affect the incorporation of proteins on solid surfaces. Much of this work is performed on artificial templates made of polymer sponges or porous materials based on alumina, mica, and porous silicon (PSi) surfaces. For example, porous silicon materials have high biocompatibility, biodegradability, and photoluminescence, which allow them to be used both as a support structure for lipid bilayers or a template to measure the electrochemical functionality of living cells grown over the surface as in vivo. The variety of these media, coupled with the complex physiological conditions present in living systems, warrant a summary and prospectus detailing which artificial systems provide the most promise for different biological conditions. This study summarizes the use of electrochemical impedance spectroscopy (EIS) data on artificial biological membranes that are closely matched with previously published biological systems using both black lipid membrane and patch clamp techniques. 相似文献
18.
Bibhas Amatya Hewang Lee Laureano D. Asico Prasad Konkalmatt Ines Armando Robin A. Felder Pedro A. Jose 《International journal of molecular sciences》2021,22(5)
The SNX-PXA-RGS-PXC subfamily of sorting nexins (SNXs) belongs to the superfamily of SNX proteins. SNXs are characterized by the presence of a common phox-homology (PX) domain, along with other functional domains that play versatile roles in cellular signaling and membrane trafficking. In addition to the PX domain, the SNX-PXA-RGS-PXC subfamily, except for SNX19, contains a unique RGS (regulators of G protein signaling) domain that serves as GTPase activating proteins (GAPs), which accelerates GTP hydrolysis on the G protein α subunit, resulting in termination of G protein-coupled receptor (GPCR) signaling. Moreover, the PX domain selectively interacts with phosphatidylinositol-3-phosphate and other phosphoinositides found in endosomal membranes, while also associating with various intracellular proteins. Although SNX19 lacks an RGS domain, all members of the SNX-PXA-RGS-PXC subfamily serve as dual regulators of receptor cargo signaling and endosomal trafficking. This review discusses the known and proposed functions of the SNX-PXA-RGS-PXC subfamily and how it participates in receptor signaling (both GPCR and non-GPCR) and endosomal-based membrane trafficking. Furthermore, we discuss the difference of this subfamily of SNXs from other subfamilies, such as SNX-BAR nexins (Bin-Amphiphysin-Rvs) that are associated with retromer or other retrieval complexes for the regulation of receptor signaling and membrane trafficking. Emerging evidence has shown that the dysregulation and malfunction of this subfamily of sorting nexins lead to various pathophysiological processes and disorders, including hypertension. 相似文献
19.
20.
The effect of proteins upon diethyl ether extraction of phospholipids from aqueous dispersions has been investigated as a model for elucidating lipid-protein interactions in the more complex membrane systems. Mixed phospholipids having a net anionic charge or purified lecithin (a zwitterion) are extracted from water dispersions into ether only after addition of salts. Basic proteins (lysozyme and cytochromec), by ionically binding phospholipids, allow extraction. Phospholipids are extracted together with the proteins in the form of neutralized lipid-protein complexes. On the other hand, lipid depleted mitochondria (a hydrophobic protein residue after acetone extraction of mitochondria), after reconstitution with phospholipids, do not allow phospholipid extraction unless salts or basic proteins also are added to the system. This observation indicates that, in reconstituted membranes, the phospholipids are largely in the bilayer form with the polar heads still charged and susceptible to ether extraction only after neutralization with salts or basic proteins. 相似文献