Preparation and properties of DNA–lipid complexes carrying pyrene and anthracene moieties

Novel DNA–lipid complexes carrying pyrene and anthracene were prepared by substituting sodium counter cations with cationic amphiphilic lipids, namely lipid(PY) and lipid(Anth), in which the actual mole ratios of phosphate to lipid were 1:1.11 and 1:1.03, respectively. DNA–lipid(PY) and DNA–lipid(Anth) complexes were soluble in common organic solvents including CHCl₃, CH₂Cl₂, methanol and ethanol, while insoluble in THF, toluene, and aqueous solutions. CD spectroscopy revealed that DNA–lipid(PY) and DNA–lipid(Anth) complexes took a predominantly double helical structure in methanol and that the helical structure was fairly stable against heating. The solution of DNA–lipid(PY) complex emitted fluorescence in 27.8% quantum yield, which were higher than that of the corresponding lipid(PY) (16.8%), while the fluorescence quantum yields of the solution of DNA–lipid(Anth) (45.4%) was lower than that of the lipid(Anth) (53.0%). The onset temperatures of weight loss of DNA–lipid(PY) and DNA–lipid(Anth) were both 220 °C according to TGA in air.     

Maximizing the Potency of siRNA Lipid Nanoparticles for Hepatic Gene Silencing In Vivo

Special (lipid) delivery: The role of the ionizable lipid pK(a) in the in?vivo delivery of siRNA by lipid nanoparticles has been studied with a large number of head group modifications to the lipids. A tight correlation between the lipid pK(a) value and silencing of the mouse FVII gene (FVII ED(50) ) was found, with an optimal pK(a) range of 6.2-6.5. The most potent cationic lipid from this study has ED(50) levels around 0.005?mg?kg(-1) in mice and less than 0.03?mg?kg(-1) in non-human primates.     

Weakly coupled lipid bilayer membranes on multistimuli-responsive poly(N-isopropylacrylamide) copolymer cushions

Polymer-cushioned lipid bilayers are frequently used to mimic the native environment of cellular membranes in respect to the extracellular matrix and intracellular structures. With the aim to actively tune lipid membrane characteristics, we pursue the approach to use temperature and pH responsive polymer thin films of poly(N-isopropylacrylamide-co-carboxyacrylamide) (PNIPAAm-co-carboxyAAM) as cushions for supported lipid bilayers. A cationic lipid bilayer composed of dioleoylphosphatidylcholine (DOPC) and dioleoyltrimethylammoniumpropane (DOTAP) (9:1) was formed on top of the polymer thin film in a drying/rehydration process. Fluorescence recovery after photobleaching (FRAP) yielded higher lipid diffusion coefficients (6.3-9.6 μm(2) s(-1)) on polymer cushions in comparison to solid glass supports (3.0-5.9 μm(2) s(-1)). No correlation of the lipid mobility was found with the swelling state of (PNIPAAm-co-carboxyAAM), which is ascribed to restrained interfacial electrostatic interactions and dispersion forces. The results revealed a minimal coupling of the lipid bilayer with the polymer cushions, and thus, bilayers supported by (PNIPAAm-co-carboxyAAM) provide interesting opportunities for unperturbed lipid diffusion combined with control of transmembrane protein mobility due to the impact of a tunable frictional drag.     

膜片电极支撑自组装双层脂膜中短杆菌肽离子通道的形成

通过一个两步程序在膜片电极尖端形成自组装双层脂膜：（1）膜片电极尖端沾取成膜液；（2）将吸附成膜液的尖端浸入电解液中，排除尖端多余的成膜液，通过电学方法监测双层脂膜的形成。将短杆菌肽通道蛋白分散在成膜液和电解质溶液中，在制备膜片电极支撑双层脂膜过程中，短杆菌肽重组到双层脂膜中形成离子通道，对通道的一般特性进行了研究，并观察到通道开放和关闭的现象。     

Formation of polyelectrolyte multilayer architectures with embedded DMPC studied in situ by neutron reflectometry

The coupling of lipid molecules to polymer components in a planar biomimetic model membrane made of a lipid bilayer (dimyristoylphosphatidylcholine) supported by polyelectrolyte multilayers is studied. The polyelectrolyte support was prepared by layer-by-layer deposition of positively charged poly(allylamine hydrochloride) (PAH) and negatively charged poly(sodium 4-styrenesulfonate) (PSS). Two polymer sample terminations were considered: positively charged (PAH-terminated) and negatively charged (PSS-terminated). Neutron reflectometry studies showed that, whereas positively charged samples did not favor the deposition of lipid, negatively charged samples allowed the deposition of a lipid bilayer with a thickness of approximately 5 nm. In the latter case, formation of polyelectrolyte layers after the deposition of the lipid layer was also possible.     

Interactions of poly(amidoamine) dendrimers with Survanta lung surfactant: the importance of lipid domains

The interaction of generation 5 (G5) and 7 (G7) poly(amidoamine) (PAMAM) dendrimers with mica-supported Survanta bilayers is studied with atomic force microscopy (AFM). In these experiments, Survanta forms distinct gel and fluid domains with differing lipid composition. Nanoscale defects are induced by the PAMAM dendrimers. The positively charged dendrimers remove lipid from the fluid domains at a significantly greater rate than for the gel domains. Dendrimer accumulation on lipid edges and terraces preceding lipid removal has been directly imaged. Immediately following lipid removal, the mica surface is clean, indicating that lipid defects are not induced by dendrimers binding to the mica substrate and displacing the lipid.     

Innate immune responses of synthetic lipid A derivatives of Neisseria meningitidis

Differences in the pattern and chemical nature of fatty acids of lipid A of Neisseria meningitides lipooligosaccharides (LOS) and Escherichia coli lipopolysaccharides (LPS) may account for differences in inflammatory properties. Furthermore, there are indications that dimeric 3-deoxy-D-manno-oct-2-ulosonic acid (KDO) moieties of LOS and LPS enhance biological activities. Heterogeneity in the structure of lipid A and possible contaminations with other inflammatory components have made it difficult to confirm these observations. To address these problems, a highly convergent approach for the synthesis of a lipid A derivative containing KDO has been developed, which relies on the ability to selectively remove or unmask in a sequential manner an isopropylidene acetal, 9-fluorenylmethoxycarbonyl (Fmoc), allyloxycarbonate (Alloc), azide, and thexyldimethylsilyl (TDS) ether. The strategy was employed for the synthesis of N. meningitidis lipid A containing KDO (3). Mouse macrophages were exposed to the synthetic compound and its parent LOS, E. coli lipid A (2), and a hybrid derivative (4) that has the asymmetrical acylation pattern of E. coli lipid A, but the shorter lipids of meningococcal lipid A. The resulting supernatants were examined for tumor necrosis factor alpha (TNF-alpha) and interferon beta (IFN-beta) production. The lipid A derivative containing KDO was much more active than lipid A alone and just slightly less active than its parent LOS, indicating that one KDO moiety is sufficient for full activity of TNF-alpha and IFN-beta induction. The lipid A of N. meningitidis was a significantly more potent inducer of TNF-alpha and IFN-beta than E. coli lipid A, which is due to a number of shorter fatty acids. The compounds did not demonstrate a bias towards a MyD88- or TRIF-dependent response.     

Ring A of nukacin ISK-1: a lipid II-binding motif for type-A(II) lantibiotic

Ring A of nukacin ISK-1, which is also present in different type-A(II) lantibiotics, resembles a lipid II-binding motif (TxS/TxD/EC, x denotes undefined residues) similar to that present in mersacidin (type-B lantibiotics), which suggests that nukacin ISK-1 binds to lipid II as a docking molecule. Results from our experiments on peptidoglycan precursor (UDP-MurNAc-pp) accumulation and peptide antagonism assays clearly indicated that nukacin ISK-1 inhibits cell-wall biosynthesis, accumulating lipid II precursor inside the cell, and the peptide activity can be repressed by lipid I and lipid II. Interaction analysis of nukacin ISK-1 and different ring A variants with lipid II revealed that nukacin ISK-1 and nukacin D13E (a more active variant) have a high affinity (K(D) = 0.17 and 0.19 μM, respectively) for lipid II, whereas nukacin D13A (a less active variant) showed a lower affinity, and nukacin C14S (a negative variant lacking the ring A structure) exhibited no interaction. Therefore, on the basis of the structural similarity and positional significance of the amino acids in this region, we concluded that nukacin ISK-1 binds lipid II via its ring A region and may lead to the inhibition of cell-wall biosynthesis.     

Cholesterol modulated antibody binding in supported lipid membranes as determined by total internal reflectance microscopy on a microfabricated high-throughput glass chip

A high-throughput microfabricated all-glass microchip, lipid biochip, was created and used to measure fluorescently tagged antibody binding to dinitrophenol (DNP) haptens in planar supported phospholipid/cholesterol lipid bilayers as a function of cholesterol-to-lipid molar ratio (X(CHOL)). Multiple parallel microchannels etched in the lipid biochip allowed simultaneous measurement of antibody binding to hapten-containing and hapten-free lipid bilayers, for a range of aqueous antibody concentrations. Specific and nonspecific antibody binding to the supported lipid bilayers was determined from the internally calibrated intensity of the surface fluorescence using total internal reflectance fluorescence (TIRF) microscopy. The TIRF intensity data of the specific antibody binding were fitted to the Langmuir isotherm and Hill equation models to determine the apparent dissociation constant K(d), the maximum fluorescence parameter F(infinity), and binding cooperativity n. As X(CHOL) increased from 0 to 0.50, K(d) exhibited a minimum of approximately 4 microM and n reached a maximum of approximately 2.2 at X(CHOL) approximately 0.20. However, F(infinity) appeared to be insensitive to the cholesterol content. The nonspecific binding fraction (NS), defined as the ratio of the TIRF intensity for hapten-free bilayers to that with hapten, showed a minimum of approximately 0.08 also at X(CHOL) approximately 0.20. The results suggest that cholesterol regulates the specific binding affinity and cooperativity, as well as suppresses nonspecific binding of aqueous antibody to a planar supported lipid bilayer surface at an optimal cholesterol content of X(CHOL) approximately 0.20. Interestingly, for X(CHOL) approximately 0.40, NS reached a maximum of approximately 0.57, suggesting significant packing defects in the lipid bilayer surface, possibly as a result of lipid domain formation as predicted by the lipid superlattice model. We conclude that cholesterol plays a significant role in regulating both specific and nonspecific antibody/antigen binding events on the lipid bilayer surface and that our lipid biochip represents a new and useful high-resolution microfluidic device for measuring lipid/protein surface binding activities in a parallel and high-throughput fashion.     

TIN ETIOPURPURIN DICHLORIDE-SENSITIZED LIPID PHOTOOXIDATION OF ERYTHROCYTE MEMBRANES

Tin(IV) etiopurpurin dichloride (SnET2 x 2Cl) is a photosensitizer which has been shown to be an effective photodynamic agent for the treatment of transplantable animal tumors in vivo. The purpose of this study was to understand the effect of SnET2 x 2Cl on membrane lipid peroxidation. When erythrocyte membranes were exposed to visible light in the presence of SnET2 x 2Cl, lipid peroxidation was observed. An accumulation of lipid hydroperoxides and an increase in lipid fluorescence were also observed. Thin layer chromatography of lipid extracts from photooxidized membrane revealed photoperoxide products derived from phospholipid. Investigations into the mechanism(s) of lipid peroxidation by SnET2 x 2Cl and light-sensitized membranes were also performed. Results indicate that singlet oxygen (1O2) plays a major role in lipid peroxidation.     

Preparation and characterization of poly(lipid)-coated, fluorophore-doped silica nanoparticles for biolabeling and cellular imaging

The fabrication, characterization, and implementation of poly(lipid)-coated, highly luminescent silica nanoparticles as fluorescent probes for labeling of cultured cells are described. The core of the probe is a sol-gel-derived silica nanoparticle, 65-100 nm in diameter, in which up to several thousand dye molecules are encapsulated (Lian, W.; et al. Anal. Biochem. 2004, 334, 135-144). The core is coated with a membrane composed of bis-sorbylphosphatidylcholine, a synthetic polymerizable lipid that is chemically cross-linked to enhance the environmental and chemical stability of the membrane relative to a fluid lipid membrane. The poly(lipid) coating has two major functions: (i) to reduce nonspecific interactions, based on the inherently biocompatible properties of the phosphorylcholine headgroup, and (ii) to permit functionalization of the particle, by doping the coating with lipids bearing chemically reactive or bioactive headgroups. Both functions are demonstrated: (i) Nonspecific adsorption of dissolved proteins to bare silica nanoparticles and of bare nanoparticles to cultured cells is significantly reduced by application of the poly(lipid) coating. (ii) Functionalization of poly(lipid)-coated nanoparticles with a biotin-conjugated lipid creates a probe that can be used to target both dissolved protein receptors as well as receptors on the membranes of cultured cells. Measurements performed on single nanoparticles bound to planar supported lipid bilayers verify that the emission intensity of these probes is significantly greater than that of single protein molecules labeled with several fluorophores.     

Measuring lipid asymmetry in planar supported bilayers by fluorescence interference contrast microscopy

There is substantial scientific and practical interest in engineering supported lipid bilayers with asymmetric lipid distributions as models for biological cell membranes. In principle, it should be possible to make asymmetric supported lipid bilayers by either the Langmuir-Blodgett/Schafer (LB/LS) or Langmuir-Blodgett/vesicle fusion (LB/VF) techniques (Kalb et al. Biochim. Biophys. Acta 1992, 1103, 307-316). However, the retention of asymmetry in biologically relevant lipid bilayers has never been experimentally examined in any of these systems. In the present work, we developed a technique that is based on fluorescence interference contrast (FLIC) microscopy to measure lipid asymmetry in supported bilayers. We compared the final degree of lipid asymmetry in LB/LS and LB/VF bilayers with and without cholesterol in liquid-ordered (l(o)) and liquid-disordered (l(d)) phases. Of five different fluorescent lipid probes that were examined, 1,2-dipalmitoyl-phosphatidylethanolamine-N-[lissamine rhodamine B] was the best for studying supported bilayers of complex composition and phase by FLIC microscopy. An asymmetrically labeled bilayer made by the LB/LS method was found to be at best 70-80% asymmetric once completed. In LB/LS bilayers of either l(o) or l(d) phase, cholesterol increased the degree of lipid mixing between the opposing monolayers. The use of a tethered polymer support for the initial monolayer did not improve lipid asymmetry in the resulting bilayer. However, asymmetric LB/VF bilayers retained nearly 100% asymmetric label, with or without the use of a tethered polymer support. Finally, lipid mixing across the center of LB/LS bilayers was found to have drastic effects on the appearance of l(d)-l(o) phase coexistence as shown by epifluorescence microscopy.     

Interactions in lipid stabilised foam films

The interaction between lipid bilayers in water has been intensively studied over the last decades. Osmotic stress was applied to evaluate the forces between two approaching lipid bilayers in aqueous solution. The force–distance relation between lipid mono- or bilayers deposited on mica sheets using a surface force apparatus (SFA) was also measured. Lipid stabilised foam films offer another possibility to study the interactions between lipid monolayers. These films can be prepared comparatively easy with very good reproducibility. Foam films consist usually of two adsorbed surfactant monolayers separated by a layer of the aqueous solution from which the film is created. Their thickness can be conveniently measured using microinterferometric techniques. Studies with foam films deliver valuable information on the interactions between lipid membranes and especially their stability and permeability. Presenting inverse black lipid membrane (BLM) foam films supply information about the properties of the lipid self-organisation in bilayers. The present paper summarises results on microscopic lipid stabilised foam films by measuring their thickness and contact angle. Most of the presented results concern foam films prepared from dispersions of the zwitterionic lipid 1,2-dimyristoyl-sn-glycero-3-phosphorylcholine (DMPC) and some of its mixtures with the anionic lipid — 1,2-dimyristoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DMPG).     

Swallow-tailed perylene derivative: a new tool for fluorescent imaging of lipid hydroperoxides

A swallow-tailed perylene derivative including a triphenylphosphine moiety was synthesized and applied to the detection and the live-cell imaging of lipid hydroperoxides. The novel probe, named Spy-LHP, reacted rapidly and quantitatively with lipid hydroperoxides to form the corresponding oxide, Spy-LHPOx, which emits extremely strong fluorescence (Phi approximately 1) in the visible range (lambda(em) = 535 nm, 574 nm). Spy-LHP was highly selective for lipid hydroperoxides, and the addition of other reactive oxygen species (ROS) including hydrogen peroxides, hydroxyl radical, superoxide anion, nitric oxide, peroxynitrite, and alkylperoxyl radical, caused no significant increase in the fluorescence intensity. The probe exhibited good localization to cellular membranes and was successfully applied to the confocal laser scanning microscopy (CLSM) imaging of lipid hydroperoxides in live J774A.1 cells, in which lipid peroxidation was proceeded by the stimulation of 2,2-azobis(2-amidinopropane)dihydrochloride (AAPH). These findings establish Spy-LHP as a promising new tool for investigating the physiology of lipid hydroperoxides.     

Quantifying growth of symmetric and asymmetric lipid bilayer domains

Here, we examine by atomic force microscopy (AFM) the kinetics and morphology of lipid domain growth during lipid phase separation by rapid thermal cooling of fully mixed two-component supported lipid bilayers. At the undercooled temperatures chosen, symmetric 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC)-rich domains favored slower reaction-limited growth whereas asymmetric galactosylceramide (GalCer)-rich domains favored faster diffusion-limited growth, indicated by shape factors and kinetic exponents. Because kinetically limited conditions could be accessed, we were able to estimate the activation energy barrier (approximately 16kT) and lateral diffusion coefficient (approximately 0.20 microm2/s) of lipid molecular addition to a growing domain. We discuss these results with respect to transition states, obstructed diffusion, and the necessity for coordinating growth in both leaflets in a symmetric lipid domain.     

Characterization and control of lipid layer fluidity in hybrid bilayer membranes

The main gel-to-liquid-crystal (LC) phase transition temperature, T(m), of the distal lipid layer in hybrid bilayer membranes (HBMs) under water was investigated using vibrational sum frequency spectroscopy (VSFS). VSFS has unique sensitivity to order/disorder transitions in the lipid acyl chains and can determine T(m) for the lipid monolayers in HBMs. We recently reported the observation that T(m) is raised and the transition width is broadened for the overlying phospholipid monolayer in HBM systems formed on densely packed crystalline self-assembled monolayers (SAMs) as compared to that of vesicles in solution. In this report, we establish that T(m) for the lipid layer of HBMs can be controlled by proper choice of the SAM underlayer. The SAM underlayer of the HBM was systematically altered by using an alkane thiol, a saturated thiolipid, a mixed SAM of a saturated lipid-pyridine disulfide, and finally a mixed SAM of an unsaturated lipid-pyridine disulfide. T(m) was measured for two different chain length saturated phosphatidylcholine lipid overlayers on these SAMs. The values obtained show that Tm of the lipid layer of HBMs is sensitive to the composition and/or packing density of molecules in the underlying SAM.     

Multicomponent cationic lipid-DNA complex formation: role of lipid mixing

Multicomponent cationic lipid-DNA complexes (lipoplexes) were prepared by adding linear DNA to mixed lipid dispersions containing two populations of binary cationic liposomes and characterized by means of small angle X-ray scattering (SAXS). Four kinds of cationic liposomes were used. The first binary lipid mixture was made of the cationic lipid (3'[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol (DC-Chol) and the neutral helper lipid dioleoylphosphocholine (DOPC) (DC-Chol/DOPC liposomes), the second one of the cationic 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and the neutral dioleoylphosphatidylethanolamine (DOPE) (DOTAP/DOPE liposomes), the third one of DC-Chol and DOPE (DC-Chol/DOPE liposomes), and the fourth one of DOTAP and DOPC (DOTAP/DOPC liposomes). Upon DNA-induced fusion of liposomes, large lipid mixing at the molecular level occurs. As a result, highly organized mixed lipoplexes spontaneously form with membrane properties intermediate between those of starting liposomes. By varying the composition of lipid dispersions, different DNA packing density regimes can also be achieved. Furthermore, occurring lipid mixing was found to induce hexagonal to lamellar phase transition in DOTAP/DOPE membranes. Molecular mechanisms underlying experimental findings are discussed.     

Molecular dynamics simulations of ternary membrane mixture: phosphatidylcholine, phosphatidic acid, and cholesterol

A ternary mixture of 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC), 1-palmitoyl-2-oleoyl phosphatidic acid (POPA), and cholesterol (CHOL) works effectively for a functional conformation of nicotinic acetylcholine receptor (nAChR) that can undergo agonist-induced conformation changes, but POPC alone can stabilize only a desensitized state of nAChR. To gain insights into the lipid mixture that has strong impact to nAChR functions, we performed more than 50 ns all atom molecular dynamic (MD) simulations at 303 K on a fully hydrated bilayer consisting of 240 POPC, 80 POPA, and 80 CHOL (3:1:1). The MD simulation revealed various interactions between different types of molecular pairs that ultimately regulated lipid organization. The heterogeneous interactions among three different constituents resulted in a broad spectrum of lipid properties, including extensive distributions of average area per lipid and varied lipid ordering as a function of lipid closeness to CHOL. Higher percentage of POPA than POPC had close association with CHOL, which coincided with relatively higher ordering of POPA molecules in their acyl chains near lipid head groups. Lower fraction of gauche dihedrals was also found in the same region of POPA. Although the CHOL molecules had the effects on the enhancement of surrounding lipid order, relatively low lipid order parameters and high fraction of gauche bonds were observed in the ternary mixture. Collectively, these results suggest that the dynamical structure of the ternary system could be determinant for a functional nAChR.     

Structure of the lipid A of Bordetella hinzii ATCC 51730

Bordetella hinzii has recently been isolated from immunocompromised human hosts. The structure of the lipid A of its endotoxin was investigated using chemical analyses, nuclear magnetic resonnance (NMR), gas liquid chromatography/mass spectrometry (GC/MS), plasma desorption mass spectrometry (PDMS) and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. The lipid A contains the classical bisphosphorylated beta-(1-->6)-linked D-glucosamine disaccharide with hydroxytetradecanoic acid (C14OH) in amide linkages. The lipid A components of B. pertussis, B. bronchiseptica, and B. parapertussis all differ in their acylation pattern but share a residue of tetradecanoyl-3-hydroxytetradecanoic acid in amide linkage at the C-2' position. However, in the B. hinzii species, the tetradecanoic acid (C14) is stoichiometrically replaced by a 2-hydroxytetradecanoic acid (2-C14OH). In the few reported examples of a hydroxylated fatty acid in this position, the substitutions were only partial. The B. hinzii lipid A differs from that of B. pertussis also by replacement of the hydroxydecanoic acid (C10OH) by hydroxydodecanoic acid (C12OH) and by the presence of a hexadecanoic acid (C16) to give a sixth fatty acid. The lipid A was heterogeneous, being composed of three major molecular species: tetra-, penta- and hexaacylated. The fatty acids in ester linkage were localized by PDMS of the native and alkali-treated lipid A. The lipid A components isolated from the O-chain-linked lipopolysaccharides (LPSs) were shown to be more acylated than those from the O-chain-free LPSs.     

Single Lipid Bilayers Constructed on Polymer Cushion Studied by Sum Frequency Generation Vibrational Spectroscopy

Planar solid supported single lipid bilayers on mica, glass, or other inorganic surfaces have been widely used as models for cell membranes. To more closely mimic the cell membrane environment, soft hydrophilic polymer cushions were introduced between the hard inorganic substrate and the lipid bilayer to completely avoid the possible substrate-lipid interactions. In this article, sum frequency generation (SFG) vibrational spectroscopy was used to examine and compare single lipid bilayers assembled on the CaF(2) prism surface and on poly (L-lactic acid) (PLLA) cushion. By using asymmetric lipid bilayers composed of a hydrogenated 1,2-dipalmitoyl-sn-glycerol-3-phosphoglycerol (DPPG) leaflet and a deuterated 1,2-dipalmitoyl-(d62)-sn-glycerol-3-phosphoglycerol (d-DPPG) leaflet, it was shown that the DPPG lipid bilayers deposited on the CaF(2) and PLLA surfaces have similar structures. SFG has also been applied to investigate molecular interactions between an antimicrobial peptide Cecropin P(1) (CP1) and the lipid bilayers on the above two different surfaces. Similar results were again obtained. This research demonstrated that the hydrophilic PLLA cushion can serve as an excellent substrate to support single lipid bilayers. We believe that it can be an important cell membrane model for future studies on transmembrane proteins, for which the possible inorganic substrate-bilayer interactions may affect the protein structure or function.