Chapter 4

the strength of alpha keratin proteins comes from?

- the cross-linking of alpha helixes by disulfide bonds

what is the predominant factor in protein stability?

- the hydrophobic effect -salt bridges can only be stable depending on the type of environment

•right-hand alpha helix

-ALPHA HELIX IS ALWAYS RIGHT HANDED!!!!!!! - R groups protruding away from the helical backbone -most common

•Secondary structure refers to a local spatial arrangement of the polypeptide backbone. •Two regular arrangements are common:

-alpha helix - beta sheet

repeated motifs..

contribute to the final fold of a protein

alpha helix contributes to

folded polypeptide chain

motif = fold =

recognizable folding pattern involving 2+ elements of secondary structures and the connection(s) can be simple, such as in a β-α-β loop (beta alpha beta) can be elaborate, such as in a β barrel (ex: green fluorescent protein)

Hair Contains Many α-Keratin Filaments

rich in hydrophobic residues: Ala, Val, Leu, Ile, Met, Phe cross-links stabilized by disulfide bonds

The backbone of a polypeptide chain can thus be pictured as a series of

rigid planes, with consecutive planes sharing a common point of rotation at - The rigid peptide bonds limit the range of conformations possible for a polypeptide chain.

Multi b-strand interactions are called

sheets. •Sheets are held together by the hydrogen bonding of amide and carbonyl groups of the peptide bond from opposite strands.

beta conformation secondary structure characteristic and example

soft, flexible filaments ex: silk- spider silk, fibroin

folding of a protein isn't always

spontaneous, sometimes you need chaperones (proteins)

ionic interactions may either be

stabilizing or destabilizing

chemical interactions stabilize native conformations:

strong disulfide (covalent) bonds are uncommon weak (noncovalent) interactions and forces are numerous hydrogen bonds hydrophobic effect ionic interactions

stability =

tendency of a protein to maintain a native conformation unfolded proteins have high conformational entropy

In antiparallel beta sheets

the H-bonded strands run in opposite directions. •Hydrogen bonds between strands are linear (stronger).

In parallel beta sheets

the H-bonded strands run in the same direction. Hydrogen bonds between strands are bent (weaker).

regulatory proteins

whereas regulatory proteins can be globular, disordered, or contain both globular and disordered segments.

repeating secondary structures (a helices and b sheets) optimize hydrogen bonding

interaction of oppositely charged groups = ion pair = salt bridge strength increases in an environment of lower dielectric constant, ε -polar aqueous solvent: ε ~ 80 -nonpolar protein interior: ε ~ 4

membrane proteins =

embedded in hydrophobic lipid membranes

The Structure of α-Keratin in Hair

- α-keratin helix is a right-handed α helix - two strands of α-keratin, oriented in parallel, wrap about each other to form a supertwisted coiled coil (= structural motif) - the alpha helixes are coiled together like a strand of rope - supertwisted helical path is left-handed but each individual alpha-keratin helix is right handed

•Unlike most organic polymers, protein molecules adopt a specific three-dimensional conformation.

- Conformation: A spatial arrangement of substituent groups that are free to assume different positions in space, without breaking any bonds, because of the freedom of bond rotation. - This structure is able to fulfill a specific biological function. - This structure is called the native fold. - There is an entropy cost to folding the protein into one specific native fold. - regions in protein that have more or less stability, also disordered regions that have no structure

how is the alpha helix stabilized?

- affected by the identities of the residues near each end - the alpha helix is not stabilized by the hydrophobic effect- this refers to the stability of the total protein structure

The Protein Data Bank (PDB):

- archive of experimentally determined three-dimensional structures - structures assigned an identifier called the PDB ID - PDB data files describe: - the spatial coordinates of each atom - information on how the structure was determined - information on its accuracy - structure visualization - software can convert atomic coordinates to an image of the molecule

•Ehlers-Danlos syndrome:

- characterized by overly flexible joints and stretchy, fragile skin. - due to collagen defects - Both conditions result from the substitution of an amino acid residue with a larger R group (such as Cys or Ser) for a single Gly residue in an α chain (a different Gly residue in each disorder).- this is defect is due to a genetic mutation

Fibrous Protein features:

- have polypeptide chains arranged in long strands or sheets. - Insoluble proteins - Give strength and/or flexibility to structures - Serve a protective or structural role - Contain polypeptide chains that generally share simple repeating element of secondary structure - H2O insoluble due to high concentrations of hydrophobic residues ex: collagen and keratin

2 rules for structural proteins:

- hydrophobic residues are in the interior - # of H bonds is maximized

The Structure of Fibroin

- main protein in silk predominantly β conformation - can stretch a lot before breaking, THINK OF A SPIDER WEB! - rich in Ala and Gly - stabilized by hydrogen bonding in beta sheets and van der Waals interactions between beta sheets

double bonds are almost always in

Trans configuration AKA DOUBLE BOND CHARACTER OF PEPTIDE BOND IS IN THE TRANS CONFIGURATION

motifs often contain conserved sequences with

distinct biological function

primary structures gives all the

info about proteins

α/β barrel =

series of β-α-β loops arranged such that the β strands form a barrel

unstructured regions are

very dynamic

•Hydrophobic effect

•The release of water molecules from the structured solvation layer around the molecule as protein folds increases the net entropy.

alpha helix

•stabilized by hydrogen bonds between nearby residues -In this structure, the polypeptide backbone is tightly wound around an imaginary axis drawn longitudinally through the middle of the helix, and the R groups of the amino acid residues protrude outward from the helical backbone - The repeating unit is a single turn of the helix -Ball-and-stick model showing the intrachain hydrogen bonds.

extended left-handed alpha helix

•theoretically less stable, not observed in proteins

•The resonance causes the peptide bonds:

•to be less reactive compared with esters, for example -resonance gives stability •to be quite rigid and nearly planar -the peptide bond is somewhat shorter than the bond in a simple amine and that the atoms associated with the peptide bond are coplanar. -This indicated a resonance or partial sharing of two pairs of electrons between the carbonyl oxygen and the amide nitrogen •partial negative charge and partial positive charge sets up a small electric dipole-. The oxygen has a partial negative charge and the hydrogen bonded to the nitrogen has a net partial positive charge, setting up a small electric dipole.

fibrous proteins =

-arranged in long strands or sheets -Fibrous proteins usually consist of a single type of secondary structure, and their tertiary structure is relatively simple. -the structures that provide support, shape, and external protection to vertebrates are made of fibrous proteins

•Globular proteins:

-fold back on each other in contrast to fibrous proteins - are soluble proteins in contrast to fibrous proteins -more compact than fibrous proteins -enzymes, transport proteins, motor proteins, regulatory proteins, immunoglobulins - globular proteins contain multiple types of secondary structure -each globular protein has a distinct structure, adapted for its biological function

globular proteins=

-folded into a spherical or globular shape - Globular proteins often contain several types of secondary structure. - most enzymes are globular proteins,

The alpha Helix: Top View

-helical wheel - The inner diameter of the helix (no side chains) is about 4-5 Å. - too small for anything to fit "inside" The outer diameter of the helix (with side chains) is 10-12 Å. (larger) Amino acids #1 and #8 align nicely on top of each other.

alpha helix secondary structure characteristic and example

-tough, insoluble protective structures of varying hardness and flexibility ex: alpha keratin of hair, feathers and nails

In summary, five types of constraints affect the stability of an α helix:

1) the intrinsic propensity of an amino acid residue to form an α helix; (2) the interactions between R groups, particularly those spaced three (or four) residues apart; (3) the bulkiness of adjacent R groups; (4) the occurrence of Pro and Gly residues; (5) interactions between amino acid residues at the ends of the helical segment and the electric dipole inherent to the α helix.

Tertiary structure is determined by amino acid sequence.

Even though protein folding is complex, some denatured proteins can spontaneously refold into their active conformation based only on the chemical properties of their constituent (primary) amino acids. - Cellular proteostasis involves numerous pathways that regulate the folding, unfolding, and degradation of proteins. Many human diseases arise from protein misfolding and defects in proteostasis.

Tertiary structure is determined by amino acid sequence.

Even though protein folding is complex, some denatured proteins can spontaneously refold into their active conformation based only on the chemical properties of their constituent amino acids. Cellular proteostasis involves numerous pathways that regulate the folding, unfolding, and degradation of proteins. Many human diseases arise from protein misfolding and defects in proteostasis.

Protein structures are stabilized by noncovalent interactions and forces.

Formation of a thermodynamically favorable structure depends on the influences of the hydrophobic effect, hydrogen bonds, ionic interactions, and van der Waals forces. Natural protein structures are constrained by peptide bonds, whose configurations can be described by the dihedral angles φ and ψ.

glycine and proline are common in beta turns

Gly and Pro residues often occur in β turns, the former because it is small and flexible, the latter because peptide bonds involving the imino nitrogen of proline readily assume the cis configuration (Fig. 4-7), a form that is particularly amenable to a tight turn

•Hydrogen bonds

Interaction of N−H and C=O of the peptide bond leads to local regular structures such as a helices and b sheets.

The position of an amino acid residue relative to its neighbors is also important.

Interactions between amino acid side chains can stabilize or destabilize the α-helical structure. For example, if a polypeptide chain has a long block of Glu residues, this segment of the chain will not form an α helix at pH 7.0. The negatively charged carboxyl groups of adjacent Glu residues repel each other so strongly that they prevent formation of the α helix. For the same reason, if there are many adjacent Lys and/or Arg residues, with positively charged R groups at pH 7.0, they also repel each other and prevent formation of the α helix. - The size and shape of Asn, Ser, Thr, and Cys residues can also destabilize an α helix if they are close together in the chain.

common beta turns in antiparallel beta sheets

Particularly common are β turns that connect the ends of two adjacent segments of an antiparallel β sheet. - The structure is a turn involving four amino acid residues, with the carbonyl oxygen of the first residue forming a hydrogen bond with the amino-group hydrogen of the fourth. The peptide groups of the central two residues do not participate in any inter-residue hydrogen bonding.

Tertiary structure describes the well-defined, three-dimensional fold adopted by a protein.

Protein structures are often built by combinatorial use of common protein folds or motifs. Quaternary structure describes the interactions between components of a multi subunit assembly.

Tertiary structure describes the well-defined, three-dimensional fold adopted by a protein.

Protein structures are often built by combinatorial use of common protein folds or motifs. - Quaternary structure describes the interactions between components of a multisubunit assembly.-Some proteins contain two or more separate polypeptide chains, or subunits, which may be identical or different. The arrangement of these protein subunits in three-dimensional complexes constitutes quaternary structure.

The three-dimensional structures of proteins can be defined.

Structural biologists use a variety of instruments and computational methods to solve biomolecular structures. The choice of method may depend on factors such as the size of the protein being studied, its properties, or the desired resolution of the final structure.

Why does the α helix form more readily than many other possible conformations?

The answer lies, in part, in its optimal use of intrahelical hydrogen bonds. The structure is stabilized by a hydrogen bond between the hydrogen atom attached to the electronegative nitrogen atom of a peptide linkage and the electronegative carbonyl oxygen atom of the fourth amino acid on the amino-terminal side of that peptide bond -Within the α helix, every peptide bond (except those close to each end of the helix) participates in such hydrogen bonding. Each successive turn of the α helix is held to adjacent turns by three to four hydrogen bonds, conferring significant stability on the overall structure. - At the ends of an α-helical segment, there are always three or four amide carbonyl or amino groups that cannot participate in this helical pattern of hydrogen bonding. These may be exposed to the surrounding solvent, where they hydrogen-bond with water, or other parts of the protein may cap the helix to provide the needed hydrogen-bonding partners.

how is secondary structure determined?

The organization around the peptide bond, paired with the identity of the R groups, determines the secondary structure of the protein.

Primary Structure: The Peptide Bond

The structure of the protein is partially dictated by the properties of the peptide bond -3 covalent bonds separate the a carbons of adjacent amino acid residues: Ca —C—N—Ca - The peptide bond is a resonance hybrid of two canonical structures

Protein segments can adopt regular secondary structures such as the α helix and the β conformation.

These structures are defined by particular values of φ and ψ and their formation is impacted by the amino acid composition of the segment. All of the φ and ψ values for a given protein structure can be visualized using a Ramachandran plot.

proline in position 2 for beta turns=

Type 1 -occurs 2x as much as type 2

glycine in position 3 for beta turns=

Type 2

secondary structure

Whereas the term "secondary structure" refers to the spatial arrangement of amino acid residues that are adjacent in a segment of a polypeptide, tertiary structure includes longer-range aspects of amino acid sequence.

The term secondary structure refers to:

any chosen segment of a polypeptide chain and describes the local spatial arrangement of its main-chain atoms, without regard to the positioning of its side chains or its relationship to other segments. -he most prominent are the α helix and β conformation; another common type is the β turn. Secondary structures without a regular pattern are sometimes referred to as undefined or as random coils

van der Waals interactions =

dipole-dipole interactions over short distances individual interactions contribute little to overall protein stability high number of interactions can be substantial

the twist of the alpha helix

ensures that critical interactions occur between an amino acid side chain and the side chain three (and sometimes four) residues away on either side of it.

•four major types of protein groups based on polypeptide chains:

fibrous globular membrane intrinsically disordered

collagen triple helix secondary structure characteristic and example

high tensile strength, without stretch ex: collagen of tendons, bone matrix

solvation layer =

highly structured shell of H2O around a hydrophobic molecule decreases when nonpolar groups cluster together decrease causes a favorable increase in net entropy hydrophobic R chains form a hydrophobic protein core

the beta sheet is primarily stabilized through?

hydrogen bonds

intrinsically disordered proteins =

lacking stable tertiary structures -Intrinsically disordered proteins can lack secondary structure entirely as well.

The native fold has a

large number of favorable interactions within the protein.

Globular proteins are composed of different

motifs folded together. - can be simple or complex

Pro and Gly occur

not commonly found in alpha helix

protein domain =

part of a polypeptide chain that is independently stable or could undergo movements as a single entity - domains are distinct structural regions that can fold and stabilize independently - domains may appear as distinct or be difficult to discern - small proteins usually have only one domain

hydrophobic effect =

predominating weak interaction

•protein family=

proteins with significant similarity in primary structure and/or tertiary structure and function are in the same protein family - protein families can be within or across organisms - ~4,000 different protein families in the PDB -strong evolutionary relationship within a family

superfamilies =

•2+ families that have little sequence similarity, but have the same major structural motif and have functional similarities -evolutionary relationship is probable

Peptide C—N Bonds Cannot Rotate Freely

•6 atoms of the peptide group lie in a single plane (planar) •partial double-bond character of C—N peptide bond prevents rotation, limiting range of conformations

alpha helix features

•= simplest arrangement, maximum number of hydrogen bonds -backbone wound around an imaginary longitudinal axis -R groups protrude out from the backbone -each helical turn = 3.6 residues, ∼5.4 Å - side chains are perpendicular to the helical z=axis - there are H bonds between backbone amides of N and N+4 amino acids

Gly helix breaker?

•Gly acts as a helix breaker because the tiny R group supports other conformations; high conformational flexibility, take up coiled structures quite different than the alpha helix.

Beta Turns Are Common in Proteins

•In globular proteins, which have a compact folded structure, some amino acid residues are in turns or loops where the polypeptide chain reverses direction - b turns: a type of protein secondary structure consisting of four amino acid residues arranged in a tight turn so that the polypeptide turns back on itself. •b turns occur frequently whenever strands in b sheets change the direction. •The 180° turn is accomplished over four amino acids. •The turn is stabilized by a hydrogen bond from a carbonyl oxygen to amide proton three residues down the sequence. Proline in position 2 or glycine in position 3 are common in b turns

3rd type of secondary structure:

•Irregular arrangement of the polypeptide chain is called the random coil.

•London dispersion

•Medium-range weak attraction between all atoms contributes significantly to the stability in the interior of the protein.

Sequence Affects Helix Stability

•Not all polypeptide sequences adopt a-helical structures. •Small hydrophobic residues such as Ala and Leu are strong helix formers. - Attractive or repulsive interactions between side chains 3 to 4 amino acids apart will affect formation.

•Two major orientations of b sheets are determined by the directionality of the strands within:

•Parallel sheets have strands that are oriented in the same direction. •Antiparallel sheets have strands that are oriented in opposite directions.

Pro, helix breaker?

•Pro acts as a helix breaker because the rotation around the N-Ca (φ-angle) bond is impossible; = introduces destabilizing kink in helix. -constraint on the formation of the α helix is the presence of Pro or Gly residues, which have the least likelihood of forming α helices. In proline, the nitrogen atom is part of a rigid ring (see Fig. 4-7), and rotation about the bond is not possible. Thus, a Pro residue introduces a destabilizing kink in an α helix. -In addition, the nitrogen atom of a Pro residue in a peptide linkage has no substituent hydrogen to participate in hydrogen bonds with other residues. For these reasons, proline is found only rarely in an α helix.

Protein Motifs Are the Basis for Protein Structural Classification

•Protein Data Bank (PDB) = 150,000+ structures archived •Structural Classification of Proteins database (SCOP2) = searches protein information in the PDB - allows a person to find relationships between protein families - proteins are classified by motifs

The Rigid Peptide Plane and the Partially Free Rotations

•Rotation around the peptide bond is not permitted due to resonance structure. •Rotation around bonds connected to the a carbon is permitted. • f (phi): angle around the a carbon—amide nitrogen bond • y (psi): angle around the a carbon—carbonyl carbon bond •In a fully extended polypeptide, both y and f are 180°- measure conformation of a peptide- By convention, ϕ and ψ are 180 degrees (or -180 degrees ) when the first and fourth atoms are farthest apart and the peptide is fully extended.

Motifs (folds) are...

•Specific arrangement of several secondary structure elements •all a helix •all b sheet •Both -A motif or fold is a recognizable folding pattern involving two or more elements of secondary structure and the connection(s) between them. •Motifs can be found as recurring structures in numerous proteins.

Amino Acid Sequence Affects Stability of the a Helix

•amino acid residues have an intrinsic propensity to form an alpha helix •interactions between R chains spaced 3-4 residues apart can stabilize or destabilize a helix -charge, size, and shape of R chains can destabilize -formation of ion pairs and hydrophobic effect can stabilize

The beta Conformation Organizes Polypeptide Chains into Sheets

•beta sheets are an extended zig zag structure - The planarity of the peptide bond and tetrahedral geometry of the a carbon create a pleated sheet-like structure. •Sheet-like arrangement of the backbone is held together by hydrogen bonds between the backbone amides in different strands. •Side chains protrude from the sheet, alternating in an up-and-down direction. -Hydrogen bonds form between backbone atoms of adjacent segments of polypeptide chain within the sheet. - The individual segments that form a β sheet are usually nearby on the polypeptide chain but can also be quite distant from each other in the linear sequence of the polypeptide; they may even be in different polypeptide chains.

Intrahelical Hydrogen Bonds

•between hydrogen atom attached to the electronegative nitrogen atom of residue n and the electronegative carbonyl oxygen atom of residue n + 4 •confers significant stability

how is tertiary structure stabilized

•by numerous weak interactions between amino acid side chains -largely hydrophobic and polar interactions -can be stabilized by covalent (disulfide) bonds •Interacting amino acids are not necessarily next to each other in the primary sequence- due to protein folding

Osteogenesis imperfecta (OI):

•characterized by abnormal bone formation in babies. - this is due to collagen defects - An inherited (genetic) bone disorder that is present at birth; also known as brittle bone disease. - A child born with OI may have soft bones that break (fracture) easily, bones that are not formed normally, and other problems. Signs and symptoms may range from mild to severe

The Structure of Collagen

•collagen is an important constituent of connective tissue: tendons, cartilage, bones, cornea of the eye. •Each collagen chain is a long Gly- and Pro-rich left-handed helix. •Three collagen chains intertwine into a right-handed superhelical triple helix. •The triple helix has higher tensile strength than a steel wire of equal cross section. •Many triple-helices assemble into a collagen fibril. - THE TRIPLE HELIX IS A UNIQUE SECONDARY STRUCTURE THAT IS DIFFERENT FROM THE ALPHA HELIX!!!!!!!!!!!

The Relationship between Protein Structure and Function

•in principle, proteins can assume an uncountable number of special arrangements, or conformations (spatial arrangements) •chemical or structural functions relate to unique three-dimensional structures

•Electrostatic interactions

•long-range strong interactions between permanently charged groups •Salt bridges (btwn oppositely charged residues) , especially those buried in the hydrophobic environment, strongly stabilize the protein.

tertiary structure =

•overall three-dimensional arrangement of all the atoms in a protein -tertiary structure= how the secondary structure is going to fold -. Amino acids that are far apart in the polypeptide sequence and are in different types of secondary structure may interact within the completely folded structure of a protein. Interacting segments of polypeptide chains are held in their characteristic tertiary positions by several kinds of weak interactions (and sometimes by covalent bonds such as disulfide cross-links) between the segments.

Myoglobin Provided Early Clues about the Complexity of Globular Protein Structure

•several structural representations of myoglobin's tertiary structure: - there are alpha helical regions, a heme group, and hydrophobic R chains

Amino Acid Residues Near the End of the a Helix Segment Affect Stability

•small electric dipoles in each peptide bond align through hydrogen bonds •negatively charged amino acids often found near the NH3+ terminus •positively charged amino acids often found near the COO- terminus - electric dipole is found throughout the alpha helix giving it an overall dipole

beta sheet

•stabilized by hydrogen bonds between adjacent segments that may not be nearby