Chapter 21: Amino Acids, Proteins, and Enzymes PDF

Chapter 21 Amino acids, Proteins, and Enzymes  Proteins Proteins serve many functions, including the following: 1. Structure: Collagen and keratin are the chief constituents of skin, bone, hair, and nails. 2. Catalysts: Virtually all reactions in living systems are...

Chapter 21 Amino acids, Proteins, and Enzymes  Proteins Proteins serve many functions, including the following: 1. Structure: Collagen and keratin are the chief constituents of skin, bone, hair, and nails. 2. Catalysts: Virtually all reactions in living systems are catalyzed by proteins called enzymes.  Proteins Proteins serve many functions, including the following: 3. Movement: Muscles are made up of proteins called myosin and actin. 4. Transport: Hemoglobin transports oxygen from the lungs to cells; other proteins transport molecules across cell membranes. 5. Hormones: Many hormones are proteins, among them insulin, oxytocin, and human growth hormone.  Proteins 6. Protection: Blood clotting involves the protein fibrinogen; the body uses proteins called antibodies to fight disease. 7. Storage: Casein in milk and ovalbumin in eggs store nutrients for newborn infants and birds. Ferritin, a protein in the liver, stores iron. 8. Regulation: Certain proteins not only control the expression of genes, but also control when gene expression takes place. Proteins  Amino Acids Amino acid: A compound that contains both an amino group and a carboxyl group.  Amino Acids Although α-amino acids are commonly written in the un- ionized form, they are more properly written in the zwitterion (internal salt) form, the form they are predominantly present at physiological pH.  Chirality of α-Amino Acids With the exception of glycine, all protein-derived amino acids have at least one stereocenter (the α-carbon) and are chiral. Glycine The vast majority of α-amino acids have the L- configuration at the α-carbon.  Chirality of α-Amino Acids A comparison of the configuration of L-alanine and D- glyceraldehyde (as Fischer projections):  Amino Acids The simplest amino acid is glycine, where R = H. The 20 Common Naturally Occurring Amino Acids  Protein-Derived α-Amino Acids Nonpolar side chains. Each ionizable group is shown in the form present in highest concentration at pH 7.0. Protein-Derived α-Amino Acids Polar side chains (at pH 7.0)  Protein-Derived α-Amino Acids Acidic and basic side chains (at pH 7.0) What is the overall charge for each amino acid?  Protein-Derived α-Amino Acids 1. For 19 of the 20, the α-amino group is primary; for proline, it is secondary. 2. With the exception of glycine, the α-carbon of each is a stereocenter. 3. Isoleucine (left) and threonine (right) contain a second stereocenter.  Ionization vs. pH An amino acid exists as a neutrally charged zwitterion at a certain pH, the isoelectric pH. The amino acid can exist in different forms, depending on the pH of the aqueous environment.  Ionization vs. pH When the pH < isoelectric pH, the carboxylate anion ____ a proton, and the amino acid has a net _____ charge.  Ionization vs. pH When the pH > isoelectric pH, the ammonium cation ____ a proton, and the amino acid has a net ____ charge.  Cysteine Amino acids have the same functional groups but differ primarily on their side chains. The properties of amino acids and their polymers, i.e. proteins, are thus determined by these side chains. The -SH (sulfhydryl) group of cysteine is easily oxidized to an -S-S- (disulfide).  Phe, Trp, and Tyr The amino acids phenylalanine, tryptophan, and tyrosine have aromatic rings on their side chains. Tryptophan is the precursor to the neurotransmitter serotonin. Bipolar disorder can be managed by controlling the levels of serotonin.  Peptides What is the product of the reaction between a carboxylic acid and amine?  Peptides In 1902, Emil Fischer proposed that proteins are long chains of amino acids joined by amide bonds. Peptide bond (peptide linkage): The special name given to the amide bond between the α-carboxyl group of one amino acid and the α-amino group of another. Peptide: A short polymer of amino acids joined by peptide bonds; they are classified by the number of amino acids in the chain. Dipeptide: A molecule containing two amino acids joined by a peptide bond.  Peptides Polypeptide: A macromolecule containing many amino acids joined by peptide bonds. Linear Polymer  Peptides Protein: A biological macromolecule containing at least 30 to 50 amino acids joined by peptide bonds. The individual amino acid units are often referred to as “residues”.  Peptides The amino acids Ala and Ser can combine with the carboxylate group of Ala reacting with the quaternary amine of Ser:  Peptides Alternatively, the carboxylate group of Ser can react with the quaternary amine of Ala: Peptides  Peptides HOW TO Draw a Dipeptide from Two Amino Acids Example Draw the structure of the dipeptide Val– Gly, and label the N-terminal and C- terminal amino acids. Step Draw the structures of the individual amino acids from left to right. Draw valine (Val) on the left. Draw glycine (Gly) on the right.  Peptides HOW TO Draw a Dipeptide from Two Amino Acids Step  Peptides HOW TO Draw a Dipeptide from Two Amino Acids Step  Peptides HOW TO Draw a Dipeptide from Two Amino Acids Step Final Answer:  Writing Peptides By convention, peptides are written from the left to right, beginning with the free -NH3+ group and ending with the free -COO- group. N-Terminus C-Terminus C-terminal amino acid: The amino acid at the end of the chain having the free -COO- group. N-terminal amino acid: The amino acid at the end of the chain having the free -NH3+ group.  Peptides Example shown : A small peptide showing the direction of the peptide chain.  Peptide Bond A peptide bond is typically written as a carbonyl group bonded to an N-H group. Linus Pauling, however, discovered that there is about 40% double bond character to the C-N bond and that a peptide bond between two amino acids is planar, which Pauling explained using the concept of resonance. C–N = 1.49Å C=N = 1.27Å Peptide Bond = 1.32Å (Partial Double Bond Character) Planar No Free Rotation Peptide Bond Levels of Structure  Levels of Structure Primary structure: The sequence of amino acids in a polypeptide chain. Read from the N-terminal amino acid to the C-terminal amino acid.  Primary Structure How important is the exact amino acid sequence? Vasopressin and oxytocin are both nonapeptides but have quite different biological functions. Vasopressin is an antidiuretic (limiting production of urine) hormone. Oxytocin affects contractions of the uterus in childbirth and the muscles of the breast that aid in the secretion of milk. The structures of vasopressin an oxytocin (next slide). Differences are shown in color.  Primary Structure Vasopressin and oxytocin are both nonapeptides but have quite different biological functions. Vasopressin is an antidiuretic (limiting production of urine) hormone. Oxytocin affects contractions of the uterus in childbirth and the muscles of the breast that aid in the secretion of milk.  Secondary Structure Secondary structure: 3D arrangement of localized regions of a protein. These regions arise due to hydrogen bonding between the N- H group of one amide with the C=O group of another. 2˚ Protein Structure: Folding of the polypeptide backbone  Secondary Structure The most common types of secondary structure are α-helix and β-pleated sheet. α-Helix: A type of secondary structure in which a section of polypeptide chain coils into a spiral, most commonly a right- handed spiral. β-Pleated sheet: A type of secondary structure in which two polypeptide chains or sections of the same polypeptide chain align parallel to each other; the chains may be parallel or antiparallel.  Secondary Structure: The α-Helix The α-Helix A single chain of protein twists into a right-handed coiled spring – a helix. The shape is maintained by intramolecular hydrogen bonds between backbone C=O and the N-H groups.  β-Pleated Sheet The β-pleated sheet structure. The backbone of two protein chains is held together by intermolecular or intramolecular hydrogen bonds.  β Sheet β-Sheet β-pleated sheets can occur between polypeptide chains (intermolecular H bonds) or in one polypeptide chain when it makes a U turn forming a hairpin structure. C N N C C N  β Sheet β-Sheet The six atoms of each peptide bond of a β-pleated sheet lie in the same plane. The C=O and N-H groups of the peptide bonds from adjacent chains point toward each other and are in the same plane so that hydrogen bonding is possible between them. All R- groups on any one chain alternate, first above, then below the plane of the sheet, etc. Anti - Parallel Parallel C N C N C N N C N C N C Purple color: side chains  Random Coil Few proteins are predominanatly α- helix or β-sheets structures. Most proteins though have only certain portions of their molecules in these conformations and the rest consists of random coil.  Secondary Structure Many globular proteins contain all three kinds of secondary structure in different parts of their molecules: α-helix, β-pleated sheet, and random coil. Schematic structure of the enzyme carboxypeptidase. The β-pleated sheet sections are shown in blue, the α-helix portions in green, and the random coils as orange strings.  Tertiary Structure Tertiary structure: the overall conformation (3-D arrangement of every atom) of a protein molecule. Complex? Includes all types of interactions. Tertiary structure is stabilized in five ways: Covalent bonds Hydrogen bonding Salt bridges/electrostatic interaction Hydrophobic interactions (London Dispersion) Metal ion coordination  Tertiary Structure Forces that stabilize tertiary structures of proteins. London dispersion Electrostatic interaction Hydrogen bonds Metal ion Covalent bonds coordination  Quaternary Structure Quaternary structure: Some proteins have more than one polypeptide chain. The quaternary structure of the protein is shape adopted when two or more folded poly-peptide chains come together into one complex.  Quaternary Structure Hemoglobin Adult hemoglobin: Made up of two identical alpha chains of 141 amino acids each, and two beta chains of 146 amino acids each. Fetal hemoglobin: Made up of two alpha chains and two gamma chains. Fetal hemoglobin has a greater affinity for oxygen than does (the alpha/beta/gamma adult hemoglobin. designations in hemoglobin have nothing to do with the α- helix and β-pleated sheets).  Proteins Proteins are divided into two types: Fibrous proteins (rod or wire-like shapes) Globular proteins (spherical or "globe-like” proteins— somewhat soluble in water as colloids)  Common fibrous proteins Collagen α keratins Vitamin C helps stabilize the chains of collagen, and, when missing, poorly formed collagen fibers result.  Common globular proteins Myoglobin has 153 Hemoglobin has 4 amino acids in 1 polypeptide chains, polypeptide chain: each carrying a heme unit. Protein Hydrolysis Hydrolysis: breaking the peptide bonds by treatment with aqueous acid, base, or certain enzymes. Pepsin in gastric juice cleaves some of the peptide bonds of large proteins. In the intestines, trypsin and chymotrypsin hydrolyze the remainder.  Denaturation Denaturation: The process of destroying the native conformation (the 3D structure) of a protein by chemical or physical means. A common method used in enzyme purification. Denaturation does not break the amide bonds that form the primary structure. That’s why it may in some cases be reversible, if the impact is small.  Denaturation Denaturing agents include: Heat: heat can disrupt hydrogen bonding; in globular proteins, it can cause unfolding of polypeptide chains with the result that coagulation and precipitation may take place. 6 M aqueous urea: Disrupts hydrogen bonding. Surface-active agents: Detergents such as sodium dodecylbenzenesulfate (SDS) disrupt hydrogen bonding. Alcohols: 70% ethanol penetrates bacteria and kills them by coagulating their proteins. It is used to sterilize skin before injections.  Denaturation Reducing agents: 2-Mercaptoethanol (HOCH2CH2SH) cleaves disulfide bonds by reducing -S-S- groups to -SH groups.  Enzymes Enzymes are proteins that catalyze biological reactions. Ribbon diagram of cytochrome c oxidase, the enzyme that directly uses oxygen during respiration.  Enzyme Catalysis Enzyme: A biological catalyst. Highly efficient catalysts: Enzymes can increase the rate of a reaction by a factor of 109 to 1020 over an uncatalyzed reaction. Specificity: Highly selective catalysts: Some enzymes catalyze the reaction of only one compound. Others are stereoselective; for example, enzymes that catalyze the reactions of only L-amino acids. Others catalyze reactions of specific types of compounds or bonds; for example, trypsin catalyzes hydrolysis of peptide bonds formed by the carboxyl groups of Lys and Arg.  Enzyme Catalysis Trypsin catalyzes the hydrolysis of peptide bonds formed by the carboxyl group of lysine and arginine.  Catalytic Power of Enzymes Enzymes provide an alternative pathway for reaction. (a) The activation energy profile for a typical reaction. (b) A comparison of the activation energy profiles for a hypothetical catalyzed and uncatalyzed reactions.  Classification of Enzymes Enzymes are commonly named after the reaction or reactions they catalyze. The names of most enzymes end with the suffix “-ase”. Example: aldehyde dehydrogenase  Cofactor A cofactor is a metal ion or an organic molecule needed for an enzyme-catalyzed reaction to occur. NAD+ is the cofactor (coenzyme) that oxidizes lactate lactate to pyruvate with the aid of the enzyme lactate dehydrogenase: Classification of Enzymes Oxidoreductases Transferases Hydrolases Lyases Ligases Isomerases  Oxidoreductase 1. Oxidoreductase: catalyze oxidation-reduction reactions. When a substrate is oxidized or reduced, a coenzyme is required, which serves are the oxidizing or reducing agent.  Transferase Transferase: catalyze transfer of a group of atoms from one molecule to another.  Hydrolase Hydrolase: catalyze hydrolysis (cleavage of bonds with water) reactions. Forward: Single bond cleavage via addition of H2O (hydrolysis) Reverse: Bond formation via the removal of H2O  Lyases Lyase: catalyze the addition of two groups to a double bond, or removal of two groups from adjacent atoms to create a double bond Not a O O O - O - hydrolysis! C H2 O C HC OPO32- C OPO32- CH2OH H2 O CH2 2-phosphoglycerate Phosphoenolpyruvate  Ligase Ligase: bond formation coupled to ATP cleavage; joining of two molecules. This process is energetically unfavorable, so the energy released in a hydrolysis reaction is used to drive the reaction.  Isomerase Isomerase: catalyze isomerization reactions. INTRAmolecular rearrangement What class does the enzyme that catalyzes this reaction belong to? A. Oxidoreductase B. Transferase C. Ligase D. Lyase E. Hydrolase F. Isomerase What class does the enzyme that catalyzes this reaction belong to? A. Oxidoreductase B. Transferase C. Ligase D. Lyase E. Hydrolase F. Isomerase What class does the enzyme that catalyzes this reaction belong to? A. Oxidoreductase B. Transferase C. Ligase D. Lyase E. Hydrolase F. Isomerase  Enzyme Terminology Substrate: The compound or compounds whose reaction an enzyme catalyzes. Active site: The specific portion of the enzyme to which a substrate binds during reaction.  Enzyme Terminology Once the reaction has occurred, the catalyst released the product(s). Two models have been proposed to explain the specificity of a substrate for an enzyme’s active site. Enzyme Specificity-the lock and key model The lock-and-key model states that the active site is a rigid cavity; to react, the substrate must exactly match the shape of the active site. Enzyme Specificity-the induced-fit model The induced-fit model states that the active site has a flexible shape, which can adjust to fit a variety of substrate shapes.  Enzyme Specificity-the induced-fit model a. Shape of the active site before binding b. Shape of the active site after binding The open cavity of the active site of the enzyme closes around the substrate for a tighter fit.  Enzyme Activity Enzyme activity: A measure of how much a reaction rate is increased over the non-catalyzed reaction. The activity of an enzyme is affected by both temperature and pH.  Allosteric Control Allosteric control is the binding of a regulator to a site on an enzyme that affects the enzyme’s ability to bind a substrate at its active site.  Terms in Enzyme Chemistry Activators: Substances that initiates or increases the activity of an enzyme. Inhibitors: Substances that makes an active enzyme less active or inactive. Inhibitor Reversible Irreversible A reversible inhibitor binds to an enzyme but then enzyme activity is restored when the inhibitor is released. An irreversible inhibitor covalently binds to an enzyme, permanently destroying its activity.  Enzyme Inhibitors Penicillin is an antibiotic that kills bacteria because it irreversibly binds to the enzyme required for the synthesis of a bacterial cell wall  Enzyme Inhibitors Inhibitor Reversible Irreversible Noncompetitive Competitive  Terms in Enzyme Chemistry Noncompetitive inhibitor: Any substance that binds to a portion of the enzyme other than the active site and thereby inhibits the activity of the enzyme. Competitive inhibitor: A substance that binds directly to the active site of an enzyme thereby preventing binding of substrate. Inhibits the action of the enzyme.  Mechanism of Inhibition The mechanism of competitive inhibition: A competitive inhibitor enters the active site, thus, preventing the substrate to enter to complex with the enzyme.  Mechanism of Inhibition Mechanism of noncompetitive inhibition: The inhibitor in this case binds itself to a site other than the active site (aka allosteric site). This changes the conformation of the active site. The substrate still binds but no catalysis takes place.  Mechanism of Action Both the lock-and-key model and the induced- fit model emphasize the shape of the active site (next slide). Just five amino acids participate in the active site in more than 65% of the enzymes studied to date. They are in order of dominance: His > Cys > Asp > Arg > Glu Four of these amino acids have either acidic or basic side chains; the fifth has a sulfhydryl group (-SH). Carboxypeptidase A Enzyme  Enzyme Regulation Feedback control (Non-covalent) Proenzymes (zymogens) (Covalent) Allosterism (Non-covalent) Protein modification (Covalent) Isoenzyme (isozymes) Proenzymes (zymogens; covalent) Zymogens (proenzymes) are an inactive form of an enzyme that can be converted to the active form when needed.  Protein Modification: Covalent The best known examples of protein modification involve phosphorylation/dephosphorylation.