Chapter 5 Appendix 133 where ␳ p is the density of the particle or macromolecule, ␳ m is the density of the medium or solution, V is the volume of the particle, and f is the frictional coeffi- cient, given by ƒ ϭ F f /v where v is the velocity of the particle and F f is the frictional drag. Nonspherical mol- ecules have larger frictional coefficients and thus smaller sedimentation coeffi- cients. The smaller the particle and the more its shape deviates from spherical, the more slowly that particle sediments in a centrifuge. Centrifugation can be used either as a preparative technique for separating and purifying macromolecules and cellular components or as an analytical technique to characterize the hydrodynamic properties of macromolecules such as proteins and nucleic acids. National Archaeological Museum, Athens, Greece/Bridgeman Art Library 6 Proteins: Secondary,Tertiary, and Quaternary Structure Nearly all biological processes involve the specialized functions of one or more pro- tein molecules. Proteins function to produce other proteins, control all aspects of cellular metabolism, regulate the movement of various molecular and ionic species across membranes, convert and store cellular energy, and carry out many other ac- tivities. Essentially all of the information required to initiate, conduct, and regulate each of these functions must be contained in the structure of the protein itself. The previous chapter described the details of protein primary structure. However, pro- teins do not normally exist as fully extended polypeptide chains but rather as com- pact structures that biochemists refer to as “folded.” The ability of a particular pro- tein to carry out its function in nature is normally determined by its overall three-dimensional shape, or conformation. This chapter reveals and elaborates upon the exquisite beauty of protein struc- tures. What will become apparent in this discussion is that the three-dimensional structure of proteins and their biological function are linked by several overarching principles: 1. Function depends on structure. 2. Structure depends both on amino acid sequence and on weak, noncovalent forces. 3. The number of protein folding patterns is very large but finite. 4. The structures of globular proteins are marginally stable. 5. Marginal stability facilitates motion. 6. Motion enables function. 6.1 What Noncovalent Interactions Stabilize the Higher Levels of Protein Structure? The amino acid sequence (primary structure) of any protein is dictated by covalent bonds, but the higher levels of structure—secondary, tertiary, and quaternary—are formed and stabilized by weak, noncovalent interactions (Figure 6.1). Hydrogen bonds, hydrophobic interactions, electrostatic bonds, and van der Waals forces are all noncovalent in nature, yet they are extremely important influences on protein con- formation. The stabilization free energies afforded by each of these interactions may be highly dependent on the local environment within the protein, but certain gener- alizations can still be made. Hydrogen Bonds Are Formed Whenever Possible Hydrogen bonds are generally made wherever possible within a given protein structure. In most protein structures that have been examined to date, component atoms of the peptide backbone tend to form hydrogen bonds with one another. Like the Greek sea god Proteus, who could assume different forms, proteins act through changes in con- formation. Proteins (from the Greek proteios, meaning “primary”) are the primary agents of biological func- tion. (“Proteus,Old Man of the Sea, Roman period mosaic, from Thessalonika,1st century a.d. National Archaeologi- cal Museum,Athens/Ancient Art and Architecture Collec- tion Ltd./Bridgeman Art Library,London/New York) Growing in size and complexity Living things, masses of atoms, DNA, protein Dancing a pattern ever more intricate. Out of the cradle onto the dry land Here it is standing Atoms with consciousness Matter with curiosity. Stands at the sea Wonders at wondering I A universe of atoms An atom in the universe. Richard P. Feynman (1918–1988) From “The Value of Science” in Edward Hutchings, Jr., ed. 1958. Frontiers of Science: A Survey. New York: Basic Books. KEY QUESTIONS 6.1 What Noncovalent Interactions Stabilize the Higher Levels of Protein Structure? 6.2 What Role Does the Amino Acid Sequence Play in Protein Structure? 6.3 What Are the Elements of Secondary Structure in Proteins, and How Are They Formed? 6.4 How Do Polypeptides Fold into Three- Dimensional Protein Structures? 6.5 How Do Protein Subunits Interact at the Quaternary Level of Protein Structure? ESSENTIAL QUESTION Linus Pauling received the Nobel Prize in Chemistry in 1954. The award cited “his research into the nature of the chemical bond and its application to the elucidation of the structure of complex substances.” Pauling pioneered the study of secondary structure in proteins. How do the forces of chemical bonding determine the formation, stability, and myriad functions of proteins? Create your own study path for this chapter with tutorials, simulations, animations, and Active Figures at www.cengage.com/ login 6.1 What Noncovalent Interactions Stabilize the Higher Levels of Protein Structure? 135 Furthermore, side chains capable of forming H bonds are usually located on the protein surface and form such bonds either with the water solvent or with other surface residues. The strengths of hydrogen bonds depend to some extent on en- vironment. The difference in energy between a side chain hydrogen bonded to wa- ter and that same side chain hydrogen bonded to another side chain is usually quite small. On the other hand, a hydrogen bond in the protein interior, away from bulk solvent, can provide substantial stabilization energy to the protein. Al- though each hydrogen bond may contribute an average of only a few kilojoules per mole in stabilization energy for the protein structure, the number of H bonds formed in the typical protein is very large. For example, in ␣-helices, the CPO and NOH groups of every interior residue participate in H bonds. The importance of H bonds in protein structure cannot be overstated. Hydrophobic Interactions Drive Protein Folding Hydrophobic “bonds,” or, more accurately, interactions, form because nonpolar side chains of amino acids and other nonpolar solutes prefer to cluster in a nonpolar en- vironment rather than to intercalate in a polar solvent such as water. The forming of hydrophobic “bonds” minimizes the interaction of nonpolar residues with water and is therefore highly favorable. Such clustering is entropically driven, and it is in fact the principal impetus for protein folding. The side chains of the amino acids in the interior or core of the protein structure are almost exclusively hydrophobic. Po- lar amino acids are much less common in the interior of a protein, but the protein surface may consist of both polar and nonpolar residues. Ionic Interactions Usually Occur on the Protein Surface Ionic interactions arise either as electrostatic attractions between opposite charges or repulsions between like charges. Chapter 4 discusses the ionization behavior of amino acids. Amino acid side chains can carry positive charges, as in the case of ly- sine, arginine, and histidine, or negative charges, as in aspartate and glutamate. In addition, the N-terminal and C-terminal residues of a protein or peptide chain usu- ally exist in ionized states and carry positive or negative charges, respectively. All of these may experience ionic interactions in a protein structure. Charged residues are normally located on the protein surface, where they may interact optimally with the water solvent. It is energetically unfavorable for an ionized residue to be located in the hydrophobic core of the protein. Ionic interactions between charged groups on a protein surface are often complicated by the presence of salts in the solution. For example, the ability of a positively charged lysine to attract a nearby negative gluta- mate may be weakened by dissolved salts such as NaCl (Figure 6.1). The Na ϩ and Cl Ϫ ions are highly mobile, compact units of charge, compared to the amino acid side chains, and thus compete effectively for charged sites on the protein. In this CH 2 CH 2 CH 2 CH 2 NH 3 Main chain Lysine H 2 O H 2 O Na + HC CO NH CO OC C O CH 2 CH 2 CH O HN NH CO Cl – Cl – Na + Main chain Glutamate +– HN FIGURE 6.1 An electrostatic interaction between the ⑀-amino group of a lysine and the ␥-carboxyl group of a glutamate.The protein is IRAK-4 kinase, an enzyme that phosphorylates other proteins (pdb id ϭ 2NRY).The interaction shown is between Lys 213 (left) and Glu 233 (right). 136 Chapter 6 Proteins: Secondary,Tertiary, and Quaternary Structure manner, ionic interactions among amino acid residues on protein surfaces may be damped out by high concentrations of salts. Nevertheless, these interactions are im- portant for protein stability. Van der Waals Interactions Are Ubiquitous Both attractive forces and repulsive forces are included in van der Waals interactions. The attractive forces are due primarily to instantaneous dipole-induced dipole inter- actions that arise because of fluctuations in the electron charge distributions of adja- cent nonbonded atoms. Individual van der Waals interactions are weak ones (with sta- bilization energies of 0.4 to 4.0 kJ/mol), but many such interactions occur in a typical protein, and by sheer force of numbers, they can represent a significant contribution to the stability of a protein. Peter Privalov and George Makhatadze have shown that, for pancreatic ribonuclease A, hen egg white lysozyme, horse heart cytochrome c, and sperm whale myoglobin, van der Waals interactions between tightly packed groups in the interior of the protein are a major contribution to protein stability. 6.2 What Role Does the Amino Acid Sequence Play in Protein Structure? It can be inferred from the first section of this chapter that many different forces work together in a delicate balance to determine the overall three-dimensional struc- ture of a protein. These forces operate both within the protein structure itself and between the protein and the water solvent. How, then, does nature dictate the man- ner of protein folding to generate the three-dimensional structure that optimizes and balances these many forces? All of the information necessary for folding the peptide chain into its “native” structure is contained in the amino acid sequence of the peptide. Just how proteins recognize and interpret the information that is stored in the amino acid sequence is not yet well understood. Certain loci along the peptide chain may act as nucleation points, which initiate folding processes that eventually lead to the correct structures. Regardless of how this process operates, it must take the protein correctly to the final native structure. Along the way, local energy- minimum states different from the native state itself must be avoided. A long-range goal of many researchers in the protein structure field is the prediction of three- dimensional conformation from the amino acid sequence. As the details of sec- ondary and tertiary structure are described in this chapter, the complexity and immensity of such a prediction will be more fully appreciated. This area is one of the greatest uncharted frontiers remaining in molecular biology. 6.3 What Are the Elements of Secondary Structure in Proteins, and How Are They Formed? Any discussion of protein folding and structure must begin with the peptide bond, the fundamental structural unit in all proteins. As we saw in Chapter 4, the resonance structures experienced by a peptide bond constrain six atoms—the oxygen, carbon, nitrogen, and hydrogen atoms of the peptide group, as well as the adjacent ␣-carbons—to lie in a plane. The resonance stabilization energy of this planar struc- ture is approximately 88 kJ/mol, and substantial energy is required to twist the structure about the CON bond. A twist of ␪ degrees involves a twist energy of 88 sin 2 ␪ kJ/mol. All Protein Structure Is Based on the Amide Plane The planarity of the peptide bond means that there are only two degrees of free- dom per residue for the peptide chain. Rotation is allowed about the bond linking the ␣-carbon and the carbon of the peptide bond and also about the bond linking 6.3 What Are the Elements of Secondary Structure in Proteins, and How Are They Formed? 137 the nitrogen of the peptide bond and the adjacent ␣-carbon. As shown in Figure 6.2, each ␣-carbon is the joining point for two planes defined by peptide bonds. The angle about the C ␣ ON bond is denoted by the Greek letter ␾ (phi), and that about the C ␣ OC o is denoted by ␺ (psi). For either of these bond angles, a value of 0° cor- responds to an orientation with the amide plane bisecting the HOC ␣ OR (side- chain) angle and a cis conformation of the main chain around the rotating bond in question (Figure 6.3). The entire path of the peptide backbone in a protein is known if the ␾ and ␺ rotation angles are all specified. Some values of ␾ and ␺ are not allowed due to steric interference between nonbonded atoms. As shown in Figure 6.3, values of ␾ ϭ 180° and ␺ ϭ 0° are not allowed because of the forbidden overlap of the NOH hydrogens. Similarly, ␾ ϭ 0° and ␺ ϭ 180° are forbidden because of unfavorable overlap between the carbonyl oxygens. G. N. Ramachandran and his co-workers in Madras, India, demonstrated that it was convenient to plot ␾ values against ␺ values to show the distribution of allowed values in a protein or in a family of proteins. A typical Ramachandran plot is shown in Fig- ure 6.4. Note the clustering of ␾ and ␺ values in a few regions of the plot. Most com- binations of ␾ and ␺ are sterically forbidden, and the corresponding regions of the Ramachandran plot are sparsely populated. The combinations that are sterically al- lowed represent the subclasses of structure described in the remainder of this section. The Alpha-Helix Is a Key Secondary Structure As noted in Chapter 5, the term secondary structure describes local conformations of the polypeptide that are stabilized by hydrogen bonds. In nearly all proteins, the hydrogen bonds that make up secondary structures involve the amide proton of one peptide group and the carbonyl oxygen of another, as shown in Figure 6.5. These structures tend to form in cooperative fashion and involve substantial por- tions of the peptide chain. When a number of hydrogen bonds form between por- tions of the peptide chain in this manner, two basic types of structures can result: ␣-helices and ␤-pleated sheets. R H Amide plane C C C C C O N N H H ␣-Carbon Side group Amide plane ␾ = 180°, ␺ =180° O Nonbonded contact radius ␾ = 0°, ␺ = 180° H C ␣ N C a H O C O H N R C a C N C a H O O H N R N H H O O H N R N H H Nonbonded contact radius ␾ = 180°, ␺ = 0° A further ␾ rotation of 120° removes the bulky carbonyl group as far as possible from the side chain ␾ = 0°, ␺ = 0° O C ␣ ␾ = –60°, ␺ = 180° H O N R H N H O O C ␣ C a C C C C C a C a C a C a C a C a C a C a C C ACTIVE FIGURE 6.3 Many of the possible conformations about an ␣-carbon between two peptide planes are forbidden because of steric crowding.Several noteworthy examples are shown here. Note: The formal IUPAC-IUB Commission on Biochemical Nomenclature convention for the definition of the torsion angles ␾ and ␺ in a polypeptide chain (Biochemistry 9:3471–3479,1970) is different from that used here, where the C ␣ atom serves as the point of reference for both rotations,but the result is the same. (Illustration: Irving Geis. Rights owned by Howard Hughes Medical Institute. Not to be reproduced without permission.) Test yourself on the concepts in this figure at www.cengage.com/login. FIGURE 6.2 The amide or peptide bond planes are joined by the tetrahedral bonds of the ␣-carbon.The rotation parameters are ␾ and ␺.The conformation shown corresponds to ␾ ϭ 180° and ␺ ϭ 180°. Note that positive values of ␾ and ␺ correspond to clockwise rotation as viewed from C ␣ . Starting from 0°, a rotation of 180° in the clockwise direction (ϩ180°) is equivalent to a rotation of 180° in the counterclockwise direction (Ϫ180°). (Illustration: Irving Geis. Rights owned by Howard Hughes Medical Institute. Not to be reproduced without permission.) 138 Chapter 6 Proteins: Secondary,Tertiary, and Quaternary Structure 180 90 –180 0 –90 ␺ (deg) –180 –90 0 90 180 ␾ (deg) ␣ II C 2 L 3 α π Antiparallel ␤-sheet Parallel ␤-sheet Collagen triple helix Right-handed ␣-helix Closed ring Left-handed ␣-helix +4 +5 –5 –4 –3 n = 2 +3 +4 +5 –5 –4 ACTIVE FIGURE 6.4 A Ramachandran diagram showing the sterically reasonable values of the angles ␾ and ␺.The shaded regions indicate particularly favorable values of these angles. Dots in purple indicate actual angles measured for 1000 residues (excluding glycine, for which a wider range of angles is permitted) in eight proteins.The lines running across the diagram (numbered ϩ5 through 2 and Ϫ5 through Ϫ3) signify the number of amino acid residues per turn of the helix; “ϩ”means right-handed helices; “Ϫ” means left-handed helices. (After Richardson, J. S., 1981.The anatomy and taxono- my of protein structure. Advances in Protein Chemistry 34:167–339.) Test yourself on the concepts in this figure at www.cengage.com/login. A DEEPER LOOK Knowing What the Right Hand and Left Hand Are Doing Certain conventions related to peptide bond angles and the “hand- edness” of biological structures are useful in any discussion of pro- tein structure. To determine the ␾ and ␺ angles between peptide planes, viewers should imagine themselves at the C ␣ carbon looking outward and should imagine starting from the ␾ ϭ 0°, ␺ ϭ 0° con- formation. From this perspective, positive values of ␾ correspond to clockwise rotations about the C ␣ ON bond of the plane that includes the adjacent NOH group. Similarly, positive values of ␺ correspond to clockwise rotations about the C ␣ OC bond of the plane that in- cludes the adjacent CPO group. Biological structures are often said to exhibit “right-hand” or “left-hand” twists. For all such structures, the sense of the twist can be ascertained by holding the structure in front of you and looking along the polymer backbone. If the twist is clockwise as one pro- ceeds outward and through the structure, it is said to be right- handed. If the twist is counterclockwise, it is said to be left-handed. 6.3 What Are the Elements of Secondary Structure in Proteins, and How Are They Formed? 139 The earliest studies of protein secondary structure were those of William Ast- bury at the University of Leeds. Astbury carried out X-ray diffraction studies on wool and observed differences between unstretched wool fibers and stretched wool fibers. He proposed that the protein structure in unstretched fibers was a he- lix (which he called the alpha form). He also proposed that stretching caused the helical structures to uncoil, yielding an extended structure (which he called the beta form). Astbury was the first to propose that hydrogen bonds between peptide groups contributed to stabilizing these structures. In 1951, Linus Pauling, Robert Corey, and their colleagues at the California In- stitute of Technology summarized a large volume of crystallographic data in a set of dimensions for polypeptide chains. (A summary of data similar to what they reported is shown in Figure 4.15.) With these data in hand, Pauling, Corey, and their colleagues proposed a new model for a helical structure in proteins, which they called the ␣-helix. The report from Caltech was of particular interest to Max Perutz in Cambridge, England, a crystallographer who was also interested in pro- tein structure. By taking into account a critical but previously ignored feature of the X-ray data, Perutz realized that the ␣-helix existed in keratin, a protein from hair, and also in several other proteins. Since then, the ␣-helix has proved to be a fundamentally important peptide structure. Several representations of the ␣-helix are shown in Figure 6.6. One turn of the helix represents 3.6 amino acid residues. (A single turn of the ␣-helix involves 13 atoms from the O to the H of the H bond. For this reason, the ␣-helix is sometimes referred to as the 3.6 13 helix.) This is in fact the feature that most confused crystallographers before the Pauling and Corey ␣-helix model. Crystallographers were so accustomed to finding twofold, threefold, sixfold, and similar integral axes in simpler molecules that the notion of a nonintegral number of units per turn was never taken seriously before Paul- ing and Corey’s work. Each amino acid residue extends 1.5 Å (0.15 nm) along the helix axis. With 3.6 residues per turn, this amounts to 3.6 ϫ 1.5 Å or 5.4 Å (0.54 nm) of travel along the helix axis per turn. This is referred to as the translation distance or the pitch of the helix. If one ignores side chains, the helix is about 6 Å in diameter. The side chains, extending outward from the core structure of the helix, are removed from R N N C C C C C C O O N N C R C O C C O FIGURE 6.5 A hydrogen bond between the backbone CPO of Ala 191 and the backbone NOH of Ser 147 in the acetylcholine-binding protein of a snail, Lymnaea stagnalis (pdb id ϭ 1I9B). Go to CengageNOW at www .cengage.com/login and click BiochemistryInteractive to explore the anatomy of the ␣-helix. 140 Chapter 6 Proteins: Secondary,Tertiary, and Quaternary Structure steric interference with the polypeptide backbone. As can be seen in Figure 6.6, each peptide carbonyl is hydrogen bonded to the peptide NOH group four residues farther up the chain. Note that all of the H bonds lie parallel to the helix axis and all of the car- bonyl groups are pointing in one direction along the helix axis while the NOH groups are pointing in the opposite direction. Recall that the entire path of the pep- tide backbone can be known if the ␾ and ␺ twist angles are specified for each residue. The ␣-helix is formed if the values of ␾ are approximately Ϫ60° and the val- ues of ␺ are in the range of Ϫ45 to Ϫ50°. Figure 6.7 shows the structures of two pro- teins that contain ␣-helical segments. The number of residues involved in a given ␣-helix varies from helix to helix and from protein to protein. On average, there are about 10 residues per helix. Myoglobin, one of the first proteins in which ␣-helices were observed, has eight stretches of ␣-helix that form a box to contain the heme prosthetic group (see Figure 5.1). As shown in Figure 6.6, all of the hydrogen bonds point in the same direction along the ␣-helix axis. Each peptide bond possesses a dipole moment that arises from the polarities of the NOH and CPO groups, and because these groups are all aligned along the helix axis, the helix itself has a substantial dipole moment, with a partial positive charge at the N-terminus and a partial negative char ge at the C-terminus (Figure 6.8). Negatively charged ligands (e.g., phosphates) frequently bind to proteins near the N-terminus of an ␣-helix. By contrast, positively charged ligands are only rarely found to bind near the C-terminus of an ␣-helix. In a typical ␣-helix of 12 (or n) residues, there are 8 (or n Ϫ 4) hydrogen bonds. As shown in Figure 6.9, the first 4 amide hydrogens and the last 4 carbonyl oxygens (d) Hydrogen bonds stabilize the helix structure. The helix can be viewed as a stacked array of peptide planes hinged at the α-carbons and approximately parallel to the helix. (a) (b) (c) α-Carbon Side group FIGURE 6.6 Four different graphic representations of the ␣-helix.(a) A stick representation with H bonds as dotted lines, as originally conceptualized in Pauling’s 1960 The Nature of the Chemical Bond. (b) Showing the arrangement of peptide planes in the helix. (Illustration: Irving Geis. Rights owned by Howard Hughes Medical Institute. Not to be repro- duced without permission.) (c) A space-filling computer graphic presentation. (d) A “ribbon structure”with an inlaid stick figure, showing how the ribbon indicates the path of the polypeptide backbone. 6.3 What Are the Elements of Secondary Structure in Proteins, and How Are They Formed? 141 ␤-Hemoglobin subunit Myohemerythrin ANIMATED FIGURE 6.7 The three- dimensional structures of two proteins that contain sub- stantial amounts of ␣-helix in their structures.The helices are represented by the regularly coiled sections of the ribbon drawings. Myohemerythrin is the oxygen-carrying protein in certain invertebrates, including Sipunculids, a phylum of marine worm. ␤-Hemoglobin subunit: pdb id ϭ 1HGA; myohemerythrin pdb id ϭ 1A7D.(Jane Richardson.) See this figure animated at www.cengage .com/login. –0.42 +0.42 –0.20 +0.20 – + Dipole moment (a) (b) C N H O FIGURE 6.8 The arrangement of NOH and CPO groups (each with an individual dipole moment) along the helix axis creates a large net dipole for the helix. Numbers indicate fractional charges on re- spective atoms. 3.6 residues C ␣8 C ␣7 C ␣5 C ␣3 C ␣2 C ␣1 C ␣4 C ␣6 O N H C ␣9 FIGURE 6.9 Four NOH groups at the N-terminal end of an ␣-helix and four CPO groups at the C-terminal end lack partners for H-bond formation.The formation of H bonds with other nearby donor and acceptor groups is referred to as helix capping. Capping may also in- volve appropriate hydrophobic interactions that accom- modate nonpolar side chains at the ends of helical segments. 142 Chapter 6 Proteins: Secondary,Tertiary, and Quaternary Structure cannot participate in helix H bonds. Also, nonpolar residues situated near the he- lix termini can be exposed to solvent. Proteins frequently compensate for these problems by helix capping—providing H-bond partners for the otherwise bare NOH and CPO groups and folding other parts of the protein to foster hydropho- bic contacts with exposed nonpolar residues at the helix termini. Careful studies of the polyamino acids, polymers in which all the amino acids are identical, have shown that certain amino acids tend to occur in ␣-helices, whereas others are less likely to be found in them. Polyleucine and polyalanine, for example, readily form ␣-helical structures. In contrast, polyaspartic acid and polyglutamic acid, which are highly negatively charged at pH 7.0, form only ran- dom structures because of strong charge repulsion between the R groups along the peptide chain. At pH 1.5 to 2.5, however, where the side chains are protonated and thus uncharged, these latter species spontaneously form ␣-helical structures. In similar fashion, polylysine is a random coil at pH values below about 11, where repulsion of positive charges prevents helix formation. At pH 12, where polylysine is a neutral peptide chain, it readily forms an ␣-helix. The tendencies of various amino acids to stabilize or destabilize ␣-helices are dif- ferent in typical proteins than in polyamino acids. The occurrence of the common amino acids in helices is summarized in Table 6.1. Notably, proline (and hydrox- yproline) act as helix breakers due to their unique structure, which fixes the value of the C ␣ ONOC bond angle. Helices can be formed from either D- or L-amino acids, but a given helix must be composed entirely of amino acids of one configu- ration. ␣-Helices cannot be formed from a mixed copolymer of D- and L-amino acids. An ␣-helix composed of D-amino acids is left-handed. The ␤-Pleated Sheet Is a Core Structure in Proteins Another type of structure commonly observed in proteins also forms because of local, cooperative formation of hydrogen bonds. That is the pleated sheet, or ␤-structure, often called the ␤-pleated sheet. This structure was also first postulated Amino Acid Helix Behavior* A Ala H (I) C Cys Variable D Asp Variable E Glu H F Phe H G Gly I (B) H His H (I) I Ile H (C) K Lys Variable L Leu H M Met H N Asn C (I) P Pro B Q Gln H (I) RArg H (I) S Ser C (B) T Thr Variable V Val Variable W Trp H (C) Y Tyr H (C) *H ϭ helix former; I ϭ indifferent; B ϭ helix breaker; C ϭ random coil; ( ) ϭ secondary tendency. TABLE 6.1 Helix-Forming and Helix-Breaking Behavior of the Amino Acids