We shall mainly discuss the membranes present in eukaryotic cells, although many of the principles described also apply to the membranes of prokaryotes. The various cellular membranes have different lipid (see below) and protein compositions. The ratio of protein to lipid in different membranes is presented in Figure 40–1, and is responsible for the many divergent functions of cellular organelles. Membranes are sheet-like enclosed structures consisting of an asymmetric lipid bilayer with distinct inner and outer surfaces or leaflets. These structures and surfaces are protein-studded, sheet-like, noncovalent assemblies that form spontaneously in water due to the amphipathic nature of lipids and the proteins contained within the membrane.
Membrane protein content is highly variable. The amount of proteins equals or exceeds the quantity of lipid in nearly all membranes. The outstanding exception is myelin, an electrical insulator found on many nerve fibers.
The Major Lipids in Mammalian Membranes Are Phospholipids, Glycosphingolipids & Cholesterol
Of the two major phospholipid classes present in membranes, phosphoglycerides are the more common and consist of a glycerol-phosphate backbone to which are attached two fatty acids in ester linkages and an alcohol (Figure 40–2). The fatty acid constituents are usually even-numbered carbon molecules, most commonly containing 16 or 18 carbons. They are unbranched and can be saturated or unsaturated with one or more double bonds. The simplest phosphoglyceride is phosphatidic acid, a 1,2-diacylglycerol 3-phosphate, a key intermediate in the formation of other phosphoglycerides (see Chapter 24). In most phosphoglycerides present in membranes, the 3-phosphate is esterified to an alcohol such as choline, ethanolamine, glycerol, inositol or serine (see Chapter 21). Phosphatidylcholine is generally the major phosphoglyceride by mass in the membranes of human cells.
A phosphoglyceride showing the fatty acids (R1 and R2), glycerol, and a phosphorylated alcohol component. Saturated fatty acids are usually attached to carbon 1 of glycerol, and unsaturated fatty acids to carbon 2. In phosphatidic acid, R3 is hydrogen.
The second major class of phospholipids comprises sphingomyelin (see Figure 21–13), a phospholipid that contains a sphingosine rather than a glycerol backbone. A fatty acid is attached by an amide linkage to the amino group of sphingosine, forming ceramide. When the primary hydroxyl group of sphingosine is esterified to phosphorylcholine, sphingomyelin is formed. As the name suggests, sphingomyelin is prominent in myelin sheaths.
The glycosphingolipids (GSLs) are sugar-containing lipids built on a backbone of ceramide. GSLs include galactosyl- and glucosyl-ceramides (cerebrosides) and the gangliosides (see structures in Chapter 21), and are mainly located in the plasma membranes of cells, displaying their sugar components to the exterior of the cell.
The most common sterol in the membranes of animal cells is cholesterol (see Chapter 21). The majority of cholesterol resides within plasma membranes, but smaller amounts are found within mitochondrial, Golgi complex, and nuclear membranes. Cholesterol intercalates among the phospholipids of the membrane, with its hydrophilic hydroxyl group at the aqueous interface and the remainder of the molecule buried within the lipid bilayer leaflet. From a nutritional viewpoint, it is important to know that cholesterol is not present in plants.
Lipids can be separated from one another and quantified by techniques such as column, thin-layer, and gas-liquid chromatography and their structures established by mass spectrometry and other techniques (see Chapter 4).
Membrane Lipids Are Amphipathic
All major lipids in membranes contain both hydrophobic and hydrophilic regions and are therefore termed amphipathic. If the hydrophobic region were separated from the rest of the molecule, it would be insoluble in water but soluble in organic solvents. Conversely, if the hydrophilic region were separated from the rest of the molecule, it would be insoluble in organic solvents but soluble in water. The amphipathic nature of a phospholipid is represented in Figure 40–3 and also Figure 21–24. Thus, the polar head groups of the phospholipids and the hydroxyl group of cholesterol interface with the aqueous environment; a similar situation applies to the sugar moieties of the GSLs (see below).
Diagrammatic representation of a phospholipid or other membrane lipid. The polar head group is hydrophilic, and the hydrocarbon tails are hydrophobic or lipophilic. The fatty acids in the tails are saturated (S) or unsaturated (U); the former are usually attached to carbon 1 of glycerol and the latter to carbon 2 (see Figure 40–2). Note the kink in the tail of the unsaturated fatty acid (U), which is important in conferring increased membrane fluidity.
Saturated fatty acids form relatively straight tails, whereas unsaturated fatty acids, which generally exist in the cis form in membranes, form “kinked” tails (Figure 40–3; see also Figures 21–1, 21–6). As the number of double bonds within the lipid side chains increase, the number of kinks in the tails increases. As a consequence, the membrane lipids become less tightly packed and the membrane more fluid. The problem caused by the presence of trans fatty acids in membrane lipids is described in Chapter 21.
Detergents are amphipathic molecules that are important in biochemistry as well as in the household. The molecular structure of a detergent is not unlike that of a phospholipid. Certain detergents are widely used to solubilize and purify membrane proteins. The hydrophobic end of the detergent binds to hydrophobic regions of the proteins, displacing most of their bound lipids. The polar end of the detergent is free, bringing the proteins into solution as detergent-protein complexes, usually also containing some residual lipids.
Membrane Lipids Form Bilayers
The amphipathic character of phospholipids suggests that the two regions of the molecule have incompatible solubilities. However, in a solvent such as water, phospholipids spontaneously organize themselves into micelles (Figure 40–4 and Figure 21–24), an assembly that thermodynamically satisfies the solubility requirements of the two chemically distinct regions of these molecules. Within the micelle the hydrophobic regions of the amphipathic phospholipids are shielded from water, while the hydrophilic polar groups are immersed in the aqueous environment. Micelles are usually relatively small in size (eg, ∼200 nm) and consequently are limited in their potential to form membranes. Detergents commonly form micelles.
Diagrammatic cross-section of a micelle. The polar head groups are bathed in water, whereas the hydrophobic hydrocarbon tails are surrounded by other hydrocarbons and thereby protected from water. Micelles are relatively small (compared with lipid bilayers) spherical structures.
Phospholipids and similar amphipathic molecules can form another structure, the bimolecular lipid bilayer, which also satisfies the thermodynamic requirements of amphipathic molecules in an aqueous environment. Bilayers are the key structures in biological membranes. Bilayers exist as sheets wherein the hydrophobic regions of the phospholipids are sequestered from the aqueous environment, while the hydrophilic, charged portions are exposed to water (Figure 40–5 and Figure 21–24). The ends or edges of the bilayer sheet can be eliminated by folding the sheet back upon itself to form an enclosed vesicle with no edges. The closed bilayer provides one of the most essential properties of membranes. The lipid bilayer is impermeable to most water-soluble molecules since such charged molecules would be insoluble in the hydrophobic core of the bilayer. The self-assembly of lipid bilayers is driven by the hydrophobic effect (see Chapter 2). When lipid molecules come together in a bilayer, the entropy of the surrounding solvent molecules increases due to the release of immobilized water.
Diagram of a section of a bilayer membrane formed from phospholipid molecules. The unsaturated fatty acid tails are kinked and lead to more spacing between the polar head groups, hence to more room for movement. This in turn results in increased membrane fluidity. (Slightly modified and reproduced, with permission, from Stryer L: Biochemistry, 2nd ed. Freeman, 1981. Copyright ©1981 by W. H. Freeman and Company.)
Two questions arise from consideration of the information described above. First, how many biologically important molecules are lipid-soluble and can therefore readily enter the cell? Gases such as oxygen, CO2, and nitrogen—small molecules with little interaction with solvents—readily diffuse through the hydrophobic regions of the membrane. The permeability coefficients of several ions and a number of other molecules in a lipid bilayer are shown in Figure 40–6. The electrolytes Na+, K+, and Cl− cross the bilayer much more slowly than water. In general, the permeability coefficients of small molecules in a lipid bilayer correlate with their solubilities in nonpolar solvents. For instance, steroids more readily traverse the lipid bilayer compared with electrolytes. The high permeability coefficient of water itself is surprising, but is partly explained by its small size and relative lack of charge. Many drugs are hydrophobic and can readily cross membranes and enter cells.
Permeability coefficients of water, some ions, and other small molecules in lipid bilayer membranes. The permeability coefficient is a measure of the ability of a molecule to diffuse across a permeability barrier. Molecules that move rapidly through a given membrane are said to have a high permeability coefficient. (Slightly modified and reproduced, with permission, from Stryer L: Biochemistry, 2nd ed. Freeman, 1981. Copyright © 1981.)
The second question concerns non-lipid-soluble molecules. How are the transmembrane concentration gradients for these molecules maintained? The answer is that membranes contain proteins, many of which span the lipid bilayer. These proteins either form channels for the movement of ions and small molecules or serve as transporters for molecules that otherwise could not readily traverse the lipid bilayer (membrane). The nature, properties, and structures of membrane channels and transporters are described below.
Membrane Proteins Are Associated With the Lipid Bilayer
Membrane phospholipids act as a solvent for membrane proteins, creating an environment in which the latter can function. As described in Chapter 5, the α-helical structure of proteins minimizes the hydrophilic character of the peptide bonds themselves. Thus, proteins can be amphipathic and form an integral part of the membrane by having hydrophilic regions protruding at the inside and outside faces of the membrane but connected by a hydrophobic region traversing the hydrophobic core of the bilayer. In fact, those portions of membrane proteins that traverse membranes do contain substantial numbers of hydrophobic amino acids and almost invariably have a high α-helical content. For most membranes, a stretch of ∼20 amino acids in an α-helical configuration will span the lipid bilayer.
It is possible to calculate whether a particular sequence of amino acids present in a protein is consistent with a transmembrane location. This can be done by consulting a table that lists the hydrophobicities of each of the 20 common amino acids and the free energy values for their transfer from the interior of a membrane to water. Hydrophobic amino acids have positive values; polar amino acids have negative values. The total free energy values for transferring successive sequences of 20 amino acids in the protein are plotted, yielding a so-called hydropathy plot. Values of over 20 kcal mol−1 are consistent with—but do not prove—the interpretation that the hydrophobic sequence is a transmembrane segment.
Another aspect of the interaction of lipids and proteins is that some proteins are anchored to one leaflet of the bilayer by covalent linkages to certain lipids; this process is termed protein lipidation. Lipidation can occur at protein termini (N- or C-) or internally. Common protein lipidation events are: C-terminal protein isoprenylation, cholesterylation and glycophosphatidylinositol (GPI; see Chapter 46); N-terminal protein myristoylation and internal cysteine S-prenylation and S-acylation. Such lipidation only occurs on a specific subset of proteins.
Different Membranes Have Different Protein Compositions
The number of different proteins in a membrane varies from less than a dozen very abundant proteins in the sarcoplasmic reticulum of muscle cells to hundreds in plasma membranes. Proteins are the major functional molecules of membranes and consist of enzymes, pumps and transporters, channels, structural components, antigens (eg, for histocompatibility), and receptors for various molecules. Because every type of membrane possesses a different complement of proteins, there is no such thing as a typical membrane structure. The enzymes associated with several different membranes are shown in Table 40–2.
TABLE 40–2Enzymatic Markers of Different Membranesa ||Download (.pdf) TABLE 40–2 Enzymatic Markers of Different Membranesa
|Membrane ||Enzyme |
|Plasma || |
|Endoplasmic reticulum ||Glucose-6-phosphatase |
|Golgi apparatus || |
| Cis ||GlcNAc transferase I |
| Medial ||Golgi mannosidase II |
| Trans ||Galactosyl transferase |
| Trans Golgi Network ||Sialyl transferase |
|Inner mitochondrial membrane ||ATP synthase |
Membranes Are Dynamic Structures
Membranes and their components are dynamic structures. Membrane lipids and proteins undergo turnover, just as they do in other compartments of the cell. Different lipids have different turnover rates, and the turnover rates of individual species of membrane proteins may vary widely. In some instances the membrane itself can turn over even more rapidly than any of its constituents. This is discussed in more detail in the section on endocytosis.
Another indicator of the dynamic nature of membranes is that a variety of studies have shown that lipids and certain proteins exhibit lateral diffusion in the plane of their membranes. Many nonmobile proteins do not exhibit lateral diffusion because they are anchored to the underlying actin cytoskeleton. By contrast, the transverse movement of lipids across the membrane (flip–flop) is extremely slow (see below) and does not appear to occur at an appreciable rate in the case of membrane proteins.
Membranes Are Asymmetric Structures
Proteins have unique orientations in membranes, making the outside surfaces different from the inside surfaces. An inside-outside asymmetry is also provided by the external location of the carbohydrates attached to membrane proteins. In addition, specific proteins are located exclusively on the outsides or insides of membranes.
There are also regional heterogeneities in membranes. Some, such as occur at the villous borders of mucosal cells, are almost macroscopically visible. Others, such as those at gap junctions, tight junctions, and synapses, occupy much smaller regions of the membrane and generate correspondingly smaller local asymmetries.
There is also inside-outside asymmetry of the phospholipids. The choline-containing phospholipids (phosphatidylcholine and sphingomyelin) are located mainly in the outer leaflet; the aminophospholipids (phosphatidylserine and phosphatidylethanolamine) are preferentially located in the inner leaflet. Obviously, if this asymmetry is to exist at all, there must be limited transverse mobility (flip-flop) of the membrane phospholipids. In fact, phospholipids in synthetic bilayers exhibit an extraordinarily slow rate of flip-flop; the half-life of the asymmetry in these synthetic bilayers is on the order of several weeks.
The mechanisms involved in the establishment of lipid asymmetry are not well understood. The enzymes involved in the synthesis of phospholipids are located on the cytoplasmic side of microsomal membrane vesicles. Translocases (flippases) exist that transfer certain phospholipids (eg, phosphatidylcholine) from the inner to the outer leaflet. Specific proteins that preferentially bind individual phospholipids also appear to be present in the two leaflets; thus lipid binding also contributes to the asymmetric distribution of specific lipid molecules. In addition, phospholipid exchange proteins recognize certain phospholipids and transfer them from one membrane (eg, the endoplasmic reticulum [ER]) to others (eg, mitochondrial and peroxisomal). A related issue is how lipids enter membranes. This has not been studied as intensively as the topic of how proteins enter membranes (see Chapter 49) and knowledge is still relatively meager. Many membrane lipids are synthesized in the ER. At least three pathways have been recognized: (1) transport from the ER in vesicles, which then transfer the contained lipids to the recipient membrane; (2) entry via direct contact of one membrane (eg, the ER) with another, facilitated by specific proteins; and (3) transport via the phospholipid exchange proteins (also known as lipid transfer proteins) mentioned above, which only exchanges lipids, but does not cause net transfer.
There is further asymmetry with regard to glycosphingolipids and glycoproteins; the sugar moieties of these molecules all protrude outward from the plasma membrane and are absent from its inner face.
Membranes Contain Integral & Peripheral Proteins
It is useful to classify membrane proteins into two types: integral and peripheral (Figure 40–7). Most membrane proteins fall into the integral class, meaning that they interact extensively with the phospholipids and require the use of detergents for their solubilization. Also, they generally span the bilayer as a bundle of α-helical transmembrane segments. Integral proteins are usually globular and are themselves amphipathic. They consist of two hydrophilic ends separated by an intervening hydrophobic region that traverses the hydrophobic core of the bilayer. As the structures of integral membrane proteins were being elucidated, it became apparent that certain ones (eg, transporter molecules, ion channels, various receptors, and G proteins) span the bilayer many times, whereas other simple membrane proteins (eg, glycophorin A) span the membrane only once (see Figures 42–4 and 52–5). Integral proteins are asymmetrically distributed across the membrane bilayer. This asymmetric orientation is conferred at the time of their insertion in the lipid bilayer during biosynthesis in the ER. The molecular mechanisms involved in insertion of proteins into membranes and the topic of membrane assembly are discussed in Chapter 49.
The fluid mosaic model of membrane structure. The membrane consists of a bimolecular lipid layer with proteins inserted in it or bound to either surface. Integral membrane proteins are firmly embedded in the lipid layers. Some of these proteins completely span the bilayer and are called transmembrane proteins, while others are embedded in either the outer or inner leaflet of the lipid bilayer. Loosely bound to the outer or inner surface of the membrane are the peripheral proteins. Many of the proteins and all the glycolipids have externally exposed oligosaccharide carbohydrate chains. (Reproduced, with permission, from Junqueira LC, Carneiro J: Basic Histology: Text & Atlas, 10th ed., McGraw-Hill, 2003.)
Peripheral proteins do not interact directly with the hydrophobic cores of the phospholipids in the bilayer and thus do not require use of detergents for their release. They are bound to the hydrophilic regions of specific integral proteins and head groups of phospholipids and can be released from them by treatment with salt solutions of high ionic strength. For example, ankyrin, a peripheral protein, is bound to the inner aspect of the integral protein “band 3” of the erythrocyte membrane. Spectrin, a cytoskeletal structure within the erythrocyte, is in turn bound to ankyrin and thereby plays an important role in maintenance of the biconcave shape of the erythrocyte.