Biochemistry, 4th Edition P29 pdf

9.1 What Are the Chemical and Physical Properties of Membranes? 243 or in the membrane and the transport of ions, sugars, and amino acids across mem- branes; and organizes and directs hundreds of cell signaling events. The Composition of Membranes Suits Their Functions Biological membranes may contain as much as 75% to 80% protein (and only 20% to 25% lipid) or as little as 15% to 20% protein. Membranes that carry out many enzyme-catalyzed reactions and transport activities (the inner mitochondrial mem- brane, chloroplast membranes, and the plasma membrane of Escherichia coli, for example) are typically richer in protein, whereas membranes that carry out fewer protein-related functions (myelin sheaths, the protective coating around neurons, for example) are richer in lipid. Cellular mechanisms adjust lipid composition to functional needs. Thus, for ex- ample, the lipid makeup of red blood cell membranes is consistent across species, whereas the lipid complement of different (specialized) membranes within a par- ticular cell type (rat liver, Figure 9.2) reflects differences of function. Plasma mem- branes are enriched in cholesterol but do not contain diphosphatidylglycerol (a) (b) (c) (d) FIGURE 9.1 Electron micrographs of several different membrane structures: (a) Plasma membrane of Menoid- ium, a protozoan. (b) Two plasma membranes from adjacent neurons in the central nervous system. (c) Golgi apparatus. (d) Many membrane structures are evident in pancreatic acinar cells. T. J. Beveridge/ Visuals Unlimited Dr. Dennis Kunkel/Visuals Unlimited © SPL/Photo Researchers, Inc. © D. W. Fawcett/Photo Researchers, Inc. Nuclear membrane 15 1333 103449 28 4 6 12 16 286 33 8 13 9 273 417 34 385 17 421771053 Plasma membrane Lysosomes Mitochondri a Phosphatidylcholine Phosphatidylethanolamine Sphingolipids Phosphatidylinositol Phosphatidylserine Cardiolipin Minor lipids Cholesterol Golgi apparatus FIGURE 9.2 The lipid composition of rat liver cell membranes, in weight percent. (Adapted from Andreoli, T.E., 1987. Membrane Physiology,2nd ed. Chapter 27, Table II,and Daum, G., 1985. Lipids of mitochondria. Biochimica et Biophysica Acta 822:1–42.) 244 Chapter 9 Membranes and Membrane Transport (cardiolipin), whereas mitochondria contain considerable amounts of cardiolipin (essential for some mitochondrial proteins) and no cholesterol. The protein com- ponents of membranes vary even more greatly than their lipid compositions. Lipids Form Ordered Structures Spontaneously in Water Monolayers and Micelles Amphipathic lipids spontaneously form a variety of struc- tures when added to aqueous solution. All these structures form in ways that minimize contact between the hydrophobic lipid chains and the aqueous milieu. For example, when small amounts of a fatty acid are added to an aqueous solution, a monolayer is formed at the air–water interface, with the polar head groups in contact with the wa- ter surface and the hydrophobic tails in contact with the air (Figure 9.3). Few lipid molecules are found as monomers in solution. Further addition of fatty acid eventually results in the formation of micelles. Micelles formed from an amphipathic lipid in water position the hydrophobic tails in the center of the lipid aggregation with the polar head groups facing outward. Amphipathic molecules that form micelles are characterized by a unique critical micelle concentration, or CMC. Below the CMC, individual lipid molecules pre- dominate. Nearly all the lipid added above the CMC, however, spontaneously forms micelles. Micelles are the preferred form of aggregation in water for detergents and soaps. Some typical CMC values are listed in Figure 9.4. Lipid Bilayers Lipid bilayers consist of back-to-back arrangements of monolayers (Figure 9.3). The nonpolar portions of the lipids face the middle of the bilayer, with the polar head groups arrayed on the bilayer surface. Phospholipid bilayers form (e) Air Water Monolayer Bilayer Normal (d) Multilamellar vesicle (b) Micelles(a) Monolayers and bilayers (c) Unilamellar vesicle Inside-out Water FIGURE 9.3 Several spontaneously formed lipid structures. Drawings of (a) monolayers and bilayers, (b) mi- celles, (c) a unilamellar vesicle, (d) a multilamellar vesicle, and (e) an electron micrograph of a multilamellar Golgi structure. David Phillips/Visuals Unlimited 9.1 What Are the Chemical and Physical Properties of Membranes? 245 rapidly and spontaneously when phospholipids are added to water, and they are sta- ble structures in aqueous solution. As opposed to micelles, which are small, self- limiting structures of a few hundred molecules, bilayers may form spontaneously over large areas (10 8 nm 2 or more). Because exposure of the edges of the bilayer to solvent is highly unfavorable, extensive bilayers normally wrap around themselves and form closed vesicles (Figure 9.3). The nature and integrity of these vesicle structures are very much dependent on the lipid composition. Phospholipids can form either uni- lamellar vesicles (with a single lipid bilayer), known as liposomes, or multilamellar vesicles. These latter structures are reminiscent of the layered structure of onions. Liposomes are highly stable structures, a consequence of the amphipathic na- ture of the phospholipid molecule. Ionic interactions between the polar head groups and water are maximized, whereas hydrophobic interactions (see Chapter 2) facilitate the association of hydrocarbon chains in the interior of the bilayer. The formation of vesicles results in a favorable increase in the entropy of the solu- tion, because the water molecules are not required to order themselves around the lipid chains. It is important to consider for a moment the physical properties of the bilayer membrane, which is the basis of vesicles and also of natural membranes. Bi- layers have a polar surface and a nonpolar core. This hydrophobic core provides a substantial barrier to ions and other polar entities. The rates of movement of such species across membranes are thus quite slow. However, this same core also pro- vides a favorable environment for nonpolar molecules and hydrophobic proteins. We will encounter numerous cases of hydrophobic molecules that interact with membranes and regulate biological functions in some way by binding to or em- bedding themselves in membranes. The Fluid Mosaic Model Describes Membrane Dynamics In 1972, S. J. Singer and G. L. Nicolson proposed the fluid mosaic model for mem- brane structure, which suggested that membranes are dynamic structures com- posed of proteins and phospholipids. In this model, the phospholipid bilayer is a fluid matrix, in essence, a two-dimensional solvent for proteins. Both lipids and pro- teins are capable of rotational and lateral movement. Singer and Nicolson also pointed out that proteins can be associated with the surface of this bilayer or embedded in the bilayer to varying degrees (Figure 9.5). Triton X-100 C C CH 3 CH 3 CH 3 (OCH 2 CH 2 ) 10 M r CMC Micelle M r 625 0.24 mM 90–95,000 Octyl glucoside H HO HOH OH H H CH 2 OH O H (CH 2 ) 7 292 25 mM C 12 E 8 (Dodecyl octaoxyethylene ether) (OCH 2 CH 2 ) 8 C 12 H 25 0.071 mM Structure 538 O CH 2 CH 3 CH 3 OH CH 3 OH FIGURE 9.4 The structures of some common detergents and their physical properties. Micelles formed by detergents can be quite large.Triton X-100, for example, typically forms micelles with a total molecular mass of 90 to 95 kD.This corresponds to approximately 150 molecules of Triton X-100 per micelle. 246 Chapter 9 Membranes and Membrane Transport They defined two classes of membrane proteins. The first, called peripheral pro- teins (or extrinsic proteins), includes those that do not penetrate the bilayer to any significant degree and are associated with the membrane by virtue of ionic interac- tions and hydrogen bonds between the membrane surface and the surface of the protein. Peripheral proteins can be dissociated from the membrane by treatment with salt solutions or by changes in pH (treatments that disrupt hydrogen bonds and ionic interactions). Integral proteins (or intrinsic proteins), in contrast, possess hydrophobic surfaces that can readily penetrate the lipid bilayer itself, as well as sur- faces that prefer contact with the aqueous medium. These proteins can either insert into the membrane or extend all the way across the membrane and expose them- selves to the aqueous solvent on both sides. Singer and Nicolson also suggested that a portion of the bilayer lipid interacts in specific ways with integral membrane pro- teins and that these interactions might be important for the function of certain membrane proteins. Because of these intimate associations with membrane lipid, integral proteins can be removed from the membrane only by agents capable of breaking up the hydrophobic interactions within the lipid bilayer itself (such as detergents and organic solvents). The fluid mosaic model became the paradigm for modern studies that have advanced our understanding of membrane structure and function. The Thickness of a Membrane Depends on Its Components Electron micro- graphs of typical cellular membranes show the thickness of the entire membrane— including lipid bilayer and embedded protein—to be 50 Å or more. Electron mi- croscopy, NMR, and X-ray and neutron diffraction measurements have shown that membrane thickness is influenced by the particular lipids and proteins in the mem- brane. The thickness of a phospholipid bilayer made from dipalmitoyl phos- phatidylcholine, measured as the phosphorus-to-phosphorus spacing, is about 37 Å, and the hydrophobic phase of such membranes is approximately 26 Å thick. Nat- ural membranes are thicker overall than simple lipid bilayers because many mem- brane proteins extend out of the bilayer significantly. Among the known membrane protein structures, there is considerable variation in the hydrophobic surface perpendicular to the membrane plane. If the hy- drophobic surface of the protein is larger or smaller than the lipid bilayer, the thick- ness of the lipid bilayer must be increased or decreased. The change in bilayer thickness due to membrane proteins can be as much as 5 Å. Integral proteins Peripheral protein Phospholipid membrane Glycolipid Oligosaccharide side chain Cholesterol FIGURE 9.5 The fluid mosaic model of membrane structure proposed by S. J. Singer and G. L. Nicolson. In this model, the lipids and proteins are assumed to be mobile; they can diffuse laterally in the plane of the mem- brane.Transverse motion may also occur, but it is much slower. 9.1 What Are the Chemical and Physical Properties of Membranes? 247 Lipid Chains May Bend and Tilt in the Membrane The long hydrocarbon chains of lipids are typically portrayed as more or less perpendicular to the membrane plane (Figure 9.3). In fact, the hydrocarbon tails of phospholipids may tilt and bend and adopt a variety of orientations. Typically, the portions of a lipid chain near the membrane surface lie most nearly perpendicular to the membrane plane, and lipid chain ordering decreases toward the end of the chain (toward the middle of the bilayer). Membranes Are Crowded with Many Different Proteins Membranes are crowded places, with a large number of proteins either embedded or associated in some way. The E. coli genome codes for more than a thousand membrane proteins. Moreover, as more membrane protein structures are determined (Figure 9.6), it has become apparent that many membrane proteins have large structures extending outside the lipid bilayer that share steric contacts and other interactions. Donald Engelman has suggested that most membranes are more crowded than first portrayed in Singer and Nicolson’s model (Figure 9.7). 200 180 160 140 120 100 80 60 40 20 0 1985 1987 1989 1991 1993 1995 1999 2001 2003 20051997 Total number of membrane protein structures solved Year FIGURE 9.6 Membrane protein structures, by year published. (Data from the Web site Membrane Proteins of Known 3D Structure at the laboratory of Stephen White, http://blanco.biomol.uci.edu/Membrane_Proteins_xtal.html, and from the Web site of Hartmut Michel, http://www.mpibp-frankfurt.mpg.de/michel/public/memprotstruct.html.) FIGURE 9.7 An updated model for membrane structure, as proposed by Donald Engelman. (Adapted from Engelman, D., 2005. Membranes are more mosaic than fluid. Nature 438:578–580.) 248 Chapter 9 Membranes and Membrane Transport 9.2 What Are the Structure and Chemistry of Membrane Proteins? Although the lipid bilayer constitutes the fundamental structural unit of all biolog- ical membranes, proteins carry out essentially all of the active functions of mem- branes. Singer and Nicolson defined peripheral proteins as globular proteins that interact with the membrane mainly through electrostatic and hydrogen-bonding interactions, and integral proteins as those that are strongly associated with the lipid bilayer. Another class of proteins not anticipated by Singer and Nicolson, the lipid- anchored proteins, is important in a variety of functions in different cells and tis- sues. These proteins associate with membranes by means of a variety of covalently linked lipid anchors. Peripheral Membrane Proteins Associate Loosely with the Membrane Peripheral proteins can bind to membranes in several ways (Figure 9.8). They may form ionic interactions and hydrogen bonds with polar head groups of membrane lipids or with other (integral) proteins, or they may interact with the nonpolar membrane core by inserting a hydrophobic loop or an amphipathic ␣-helix. Exam- ples of each of these interaction types are shown in Figure 9.9. Integral Membrane Proteins Are Firmly Anchored in the Membrane Hundreds of structures of integral membrane proteins are now available in the Pro- tein Data Bank, and the number of membrane protein structures is doubling about every 3 years. The known structures show a surprising diversity, but in all cases the ؉ ؊ ؉ ؊ Association with integral protein Hydrophobic loop Amphipathic ␣-helix Ionic and H-bond interactions FIGURE 9.8 Four possible modes for the binding of peripheral membrane proteins. FIGURE 9.9 Models for membrane association of peripheral proteins. (a) Bee venom phospholipase A 2 (pdb id ϭ 1POC), (b) p40 phox PX domain of NADH oxidase (pdb id ϭ 1H6H), and (c) PH domain of phospholipase C␦ (pdb id ϭ 1MAI). (a) (b) (c) 9.2 What Are the Structure and Chemistry of Membrane Proteins? 249 portions of the protein in contact with the nonpolar core of the lipid bilayer are dominated by ␣-helices or ␤-sheets, because these secondary structures neutralize the highly polar NOH and CPO functions of the peptide backbone through H- bond formation. Proteins with a Single Transmembrane Segment In proteins that are anchored by a single hydrophobic segment, that segment typically takes the form of an ␣-helix. One of the best examples is glycophorin. Most of glycophorin’s mass is ori- ented on the outside surface of the red blood cell, exposed to the aqueous milieu (Figure 9.10). Hydrophilic oligosaccharide units are attached to this extracellular domain. These oligosaccharide groups constitute the ABO and MN blood group antigenic specificities of the red cell. Glycophorin has a total molecular weight of about 31,000 and is approximately 40% protein and 60% carbohydrate. The gly- cophorin primary structure consists of a segment of 19 hydrophobic amino acid residues with a short hydrophilic sequence on one end and a longer hydrophilic sequence on the other end. The 19-residue sequence is just the right length to span the cell membrane if it is coiled in the shape of an ␣-helix. Monoamine oxidase from the mitochondrial outer membrane is another typical single transmembrane–segment protein (Figure 9.11); this enzyme is the target for many antidepressant and neuroprotective drugs. Each monomer of the dimeric protein binds to the membrane through a C-terminal transmembrane ␣-helix. Residues in two loops (Pro-109 and Ile-110 in the 99–112 loop and Phe-481, Leu- 482, Leu-486, and Pro-487 in the 481–488 loop) also provide nonpolar residues that participate in membrane binding. Approximately 10% to 30% of transmembrane proteins have a single helical transmembrane segment. In animals, many of these function as cell surface recep- tors for extracellular signaling molecules or as recognition sites that allow the im- mune system to recognize and distinguish cells of the host organism from invading Leu Ser ThrSer Thr Glu Gly Val Ala Met His Thr Thr Thr Ser SerVal Ser Ser LysSerTyrIleSerSerGlnThrAsn Asp Thr Lys His Arg Asp Thr Tyr Ala Ala Thr Pro Arg Ala His Glu Val Ser Glu Ile Ser Val Arg Thr Ile Ser Tyr Gly Ile Arg Arg Leu Ile Lys Lys Ser Pro Ser Asp Val Lys Pro Leu Leu Leu Ile Thr Ile Leu Thr Ile Glu Glu Pro Ser Phe His His Ala Leu Gln Val Arg Glu Gly Thr Glu Glu Glu Pro Pro Tyr Val Gly Ala Met Val Gly Ile Val Gly Phe Ile Pro Ser Pro Asp Thr Asp Val Pro Leu Ser Ser Val Ile Glu Asn Glu Pro Thr Ser Asp Gln COO – Glu H 3 + N — 10 20 30 40 50 60 70 100 110 120 90 130 Outside Inside Carbohydrate FIGURE 9.10 Glycophorin A spans the membrane of the human erythrocyte via a single ␣-helical transmem- brane segment.The C-terminus of the peptide faces the cytosol of the erythrocyte; the N-terminal domain is extracellular. Points of attachment of carbohydrate groups are indicated by triangles. 250 Chapter 9 Membranes and Membrane Transport foreign cells or viruses. The proteins that represent the major transplantation antigens H2 in mice (Figure 9.11) and human leukocyte associated (HLA) proteins in humans are members of this class. Other such proteins include the surface immunoglobulin recep- tors on B lymphocytes and the spike proteins of many membrane viruses. The function of many of these proteins depends primarily on their extracellular domain; thus, the segment facing the intracellular surface is often a shorter one. Proteins with Multiple Transmembrane Segments Most integral transmembrane proteins cross the lipid bilayer more than once. These multi-spanning membrane proteins typically have 2 to 12 transmembrane segments, and they carry out a vari- ety of cellular functions (Figure 9.12). A well-characterized example of such a pro- tein is bacteriorhodopsin (Figure 9.13), which clusters in purple patches in the membrane of the archaeon Halobacterium halobium. The name Halobacterium refers to the fact that this prokaryote thrives in solutions having high concentrations of (b)(a) Outside Inside Outside Inside FIGURE 9.11 (a) Major histocompatibility antigen HLA-A2 (pdb id ϭ 1JF1) and (b) monoamine oxidase (pdb id ϭ 1GOS) are membrane-associated proteins with a single transmembrane helical segment. 90 100 80 70 60 50 40 30 20 10 0 21817161514131211109865437 Number of proteins Number of transmembrane helices FIGURE 9.12 Most membrane proteins possess 2 to 12 transmembrane segments.Those involved in transport functions have between 6 and 12 transmembrane segments.(Adapted from von Heijne, G.,2006. Membrane-protein topology. Nature Reviews Molecular Cell Biology 7:909–918.) 9.2 What Are the Structure and Chemistry of Membrane Proteins? 251 sodium chloride, such as the salt ponds of San Francisco Bay. Halobacterium carries out a light-driven proton transport by means of bacteriorhodopsin, named in ref- erence to its spectral similarities to rhodopsin in the rod outer segments of the mammalian retina. The amino acid sequence of bacteriorhodopsin contains seven different segments, each about 20 nonpolar residues in length—just the right size for an ␣-helix that could span a bilayer membrane. (Twenty residues times 1.5 Å per residue equals 30 Å.) Bacteriorhodopsin clusters in symmetric, repeating arrays in the purple mem- brane patches of Halobacterium, and it was this orderly, repeating arrangement of proteins in the membrane that enabled Nigel Unwin and Richard Henderson in 1975 to determine the bacteriorhodopsin structure. The polypeptide chain crosses the membrane seven times, in seven ␣-helical segments, with very little of the pro- tein exposed to the aqueous milieu. The bacteriorhodopsin structure became a model of globular membrane protein structure. Many other integral membrane proteins contain numerous hydrophobic sequences that, like those of bacterio- rhodopsin, form ␣-helical transmembrane segments. Membrane Protein Topology Can Be Revealed by Hydropathy Plots The topol- ogy of a membrane protein is a specification of the number of transmembrane seg- ments and their orientation across the membrane. The topology of a transmem- brane helical protein can be revealed by a hydropathy plot based on its amino acid sequence. If a measure of hydrophobicity is assigned to each amino acid (Table 9.1), then the overall hydrophobicity of a segment of a polypeptide chain can be es- timated. The hydropathy index for any segment is an average of the hydrophobic- ity values for its residues. The hydropathy index can be calculated at any residue in a sequence by averag- ing the hydrophobicity values for a segment surrounding that residue. Typically, segment sizes for such calculations can be 7 to 21 residues. With a 7-residue seg- ment size, the calculation of hydropathy index at residue 10 would average the val- ues for residues 7 through 13. The calculation for a 21-residue segment around residue 100 would include residues 90 to 110. A polypeptide segment approxi- mately 20 residues long with a high hydropathy index is likely to be an ␣-helical transmembrane segment. A hydropathy plot for glycophorin (Figure 9.14a) reveals a single region of high hydropathy index between residues 73 and 93, the location of the ␣-helical segment in this transmembrane protein (Figure 9.10). A hydropa- thy plot for rhodopsin (Figure 9.14b) reveals the locations of its seven ␣-helical transmembrane segments. Rhodopsin, the light-absorbing pigment protein of the eye, is a member of the G-protein–coupled receptor (GPCR) family of membrane proteins (see Chapter 32). Proline Residues Can Bend a Transmembrane ␣-Helix Transmembrane ␣-helices often contain distortions and “kinks”—more so than for water-soluble proteins. As more integral membrane protein structures have been determined, it has become clear that most transmembrane ␣-helices contain significant distortions from ideal helix geometry. Helix distortions may have evolved in membrane proteins because (1) helices, even distorted ones, are highly stable in the membrane environment, and (2) helix distortions may be one way to create structural diversity from the sim- ple helix building blocks of most membrane proteins. About 60% of known membrane helix distortions are kinks at proline residues (Figure 9.13). Proline distorts the ideal ␣-helical geometry because of steric conflict with the preceding residue and because of the loss of a backbone H bond. Proline- induced kinks create weak points in the helix, which may facilitate movements re- quired for transmembrane transport channels. Amino Acids Have Preferred Locations in Transmembrane Helices Transmem- brane protein sequences and structures are adapted to the transition from water on one side of the membrane, to the hydrocarbon core of the membrane, and then to water on the other side of the membrane. The amino acids that make up trans- FIGURE 9.13 Bacteriorhodopsin is composed of seven transmembrane ␣-helical segments connected by short loops (pdb id ϭ 1M0M). Nearly all of this protein is embedded in the membrane. Only the short loops con- necting helices are exposed to solvent. A retinal chro- mophore (a light-absorbing molecule, shown in blue) lies approximately parallel to the membrane and between the helical segments. A proline residue (red) induces a kink in one of the helical segments (green). Side Chain Hydropathy Index Isoleucine 4.5 Valine 4.2 Leucine 3.8 Phenylalanine 2.8 Cysteine 2.5 Methionine 1.9 Alanine 1.8 Glycine Ϫ0.4 Threonine Ϫ0.7 Serine Ϫ0.8 Tryptophan Ϫ0.9 Tyrosine Ϫ1.3 Proline Ϫ1.6 Histidine Ϫ3.2 Glutamic acid Ϫ3.5 Glutamine Ϫ3.5 Aspartic acid Ϫ3.5 Asparagine Ϫ3.5 Lysine Ϫ3.9 Arginine Ϫ4.5 *From Kyte, J., and Doolittle, R., 1982. A simple method for dis- playing the hydropathic character of a protein. Journal of Molecular Biology 157:105–132. Hydropathy Scale for Amino Acid Side Chains in Proteins* Go to CengageNOW at www.cengage.com/login and click BiochemistryInteractive to explore the structure of the bacteriorhodopsin. TABLE 9.1 252 Chapter 9 Membranes and Membrane Transport membrane segments reflect these transitions. Hydrophobic amino acids (Ala, Val, Leu, Ile, and Phe) are found most often in the hydrocarbon interior, where charged and polar amino acid almost never reside (Figure 9.15b). Charged residues (Figure 9.15a) occur commonly at the lipid-water interface, but positively charged residues are found more often on the cytoplasmic face of transmembrane proteins. Gunnar von Heijne has termed this the “positive inside rule.” Tryptophan, histidine, and tyrosine are special cases (Figure 9.15c). These residues have a mixed character, with nonpolar aromatic rings that also contain polar parts (the ring NOH of Trp and the substituent OOH of Tyr). As such, Trp and Tyr are found commonly at the lipid–water interface of transmembrane proteins. The amino acids Lys and Arg frequently behave in novel ways at the lipid–water interface. Both of these residues possess long aliphatic side chains with positively charged groups at the end. In many membrane proteins, the aliphatic chain of Lys or Arg is associated with the hydrophobic portion of the bilayer, with the positively 0 –2 4 2 12010080 Hydrophobic transmembrane segment 604020 Hydropathy index Residue number 0 –2 2 4 300250200150 1 2 3 4 5 6 7 10050 Hydropathy index Residue number (a) (b) FIGURE 9.14 Hydropathy plots for (a) glycophorin and (b) rhodopsin. Hydropathy index is plotted versus residue number. At each position in the polypeptide chain, the average of hydropathy indices for a certain number of adjacent residues (eight, in this case) is calculated and plotted on the y-index, and the number of the residue in the middle of this “window” is shown on the x-axis. (b) Ϫ45 Distance from membrane center (Å) Energy Ϫ15 15 45 (c) Ϫ45 Distance from membrane center (Å) Energy Ϫ15 15 45 (a) Ϫ45 Distance from membrane center (Å) Energy Ϫ15 15 45000 FIGURE 9.15 Amino acids have distinct preferences for different parts of the membrane.The graphs show rela- tive stabilization energies as a function of location in the membrane for (a) Arg, Asp, Glu, Lys, Asn, Gln, and Pro; (b) Ala,Gly, Ile, Leu, Met, Phe, and Val; and (c) His,Tyr, and Trp.Polar and charged residues are less stable in the membrane interior, whereas nonpolar residues tend to be more stable in the membrane interior.The stability profiles for His,Tyr, and Trp are more complex. (Adapted from von Heijne, G., 2006. Membrane-protein topology. Nature Reviews Molecular and Cell Biology 7:909–918.)