Biochem Chapter 5 notes
Biochem Chapter 5 notes 87222 - BCHM 3050 - 002
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Biochem Notes Chapter 5: Amino Acids, Peptides, and Proteins proteins can be distinguished based on their number of amino acids (amino acid residues), their overall amino acyl composition, and their amino acyl sequence Polypeptides: molecules with molecular weights ranging from several thousand to several million daltons peptides: low molecular weight, consisting of fewer than 50 amino acids protein: describes molecules with more than 50 amino acids, used interchangeably with polypeptide 5.1: Amino Acids there are 20 standard amino acids; they contain a central carbon atom (alpha carbon) to which an amino group, a carboxylate group, a hydrogen group, and an R group (side chain) are attached nonstandard amino acids consist of amino acid residue s that have been chemically modified after incorporation into a polypeptide or amino acids that occur in living organisms but are not found in proteins At pH of 7, the carboxyl group of an amino acid is in its conjugate base form (COO) and the amino group is in its conjugate acid form (NH3+) amphoteric: can act as an acid or a base Zwitterions: molecules that have a negative and a positive charge on different atoms Amino acid classes: o Nonpolar: contain mostly hydrocarbon R groups that do not bear a positive or negative charge; interact poorly with water (hydrophobic); two types of hydrocarbon side chains aromatic (contain cyclic structures that constitute a class of unsaturated hydrocarbons) and aliphatic (non aromatic hydrocarbons such as methane and cyclohexane); Phenylalanine, tryptophan, glycine, alanine, valine, leucine, isoleucine, proline, and cystine o polar: have functional groups capable of hydrogen bonding; hydrophilic and easily interact with water; serine, threonine, tyrosine, asparagine, glutamine o acidic: have a sidechain with carboxylate groups; are negatively charged at physiological pH; Aspartate and glutamate o basic: have positive charges at physiological pH; lysine, arginine, and histidine o amino acids are classified according to their capacity to interact with water; this criterion may be used to distinguish the four classes o Biologically Active Amino Acids: 1. Neurotransmitters: several alphaamino acids or their derivatives act as chemical messengers; Hormones: chemical signal molecules produced in one cell that regulates the function of other cells 2. amino acids are precursors of a variety of complex nitrogencontaining molecules; includes components of nucleotides and nucleic acids, heme (ironcontaining organic group required for the biological activity of several important proteins), and chlorophyll 3. Several standard and nonstandard amino acids act as metabolic intermediates Modified Amino Acids in Proteins: several proteins contain amino acid derivatives that are formed after a polypeptide chain has been synthesized Amino Acid Stereoisomers: because the alphacarbon of 19 out of the 20 amino acids have four different groups attached, they are known as asymmetric, or chiral, carbons; molecules with chiral carbons can exist as stereoisomers (molecules that differ only in the spatial arrangement of their atoms); two isomers that are mirror images of each other are enantiomers; optical isomers are when the light waves of a planepolarized light vibrate in only one place molecules with asymmetric or chiral carbon atoms differ only in the spatial arrangement of the atoms attached to the carbon the mirrorimage forms of a molecule are called enantiomers most asymmetric molecules in living organisms occur in only one stereoisometric form Titration of amino acids: because amino acids contain ionizable groups, the predominant ionic form of these molecules in solution depends on the pH; Isoelectric point (pI): is the pH at which the carboxyl group has lost its proton and has no net charge, electrically neutral can be calculated by pI= (pK1 + pK2) 2; as titration continues, the ammonium group loses its proton leaving an uncharged amino group giving the molecule a negative charge When amino acids are incorporated in polypeptides, the aminoand carboxylgroup lose their charges; consequently, except for the N and the C terminal residues, all the ionizable groups of proteins are the side chain groups of seven amino acids: histidine, lysine, arginine, aspartate, glutamate, cysteine, tyrosine Amino Acid Reactions: o Peptide bond formation: peptide bonds are amide linkages formed when the unshared electron pair of the alphaamino nitrogen atom of one amino acid attacks the acarboxyl carbon of another in a nucleophilic acyl substitution reaction o have an amino acid sequence: the order in which the amino acids are linked together o Cysteine Oxidation: the sulfahydryl group of cysteine is highly reactive; oxidation of two molecules of cysteine form cystine, a molecule with a disulfide bond; when two cysteine residues form such a bond, it is referred to as a disulfide bridge that help stabilize polypeptides and proteins o Schiff Base Formation: molecules that possess primary amine groups can reversibly react with carbonyl groups; the imine products are known as Schiff bases; the most important examples of Schiff base formation in biochem occur in amino acid metabolism aldimines formed by the reversible reaction of an amino group with an aldehyde are the intermediates o Polypeptides are polymers composed of amino acids linked by peptide bonds; the order of the amino acids in the polypeptide is called amino acid sequence o disulfide bridges, formed by the oxidation of cysteine residues, are an important structural element in polypeptides and proteins o schiff bases are imines that form when amine groups react reversibly with carbonyl groups 5.2 PEPTIDES the tripeptide glutathione contains an unusual yamide bond; found in almost all organisms, it is involved in protein and DNA synthesis, drug and environmental toxin metabolism, amino acid transport and other biological processes; an important intracellular antioxidant homeostasis: process to maintain a stable internal environment vasopressin: antidiuretic hormone; affects blood volume; contains nine amino acid residues; synthesized in the hypothalamus; released in response to low blood pressure or high Na+ in the blood oxytocin: stimulates the ejection of milk by the mammary glands atrial natriuretic factor (ANF): peptide produced by the heart in response to stretching; also stimulates the production of dilute urine (opposite of vasopressin) Although small in comparison to larger protein molecules, peptides have significant biological activity; they are involved in a variety of signal transduction processes 5.3 PROTEINS have the most diverse functions: 1. catalysis: enzymes are proteins that accelerate chemical reactions; the can perform under mild conditions of pH and temperature because they can induce or stabilize strained reaction intermediates 2. structure: provide structural support; have specialized properties (EX: collagen, fibroin have mechanical strength; elastin is in the muscles that have to be elastic to function) 3. movement: proteins are involved in all cell movements 4. defense: keratin (found in skin cells) protects the organism against mechanical and chemical injury; antibodies; blood clotting proteins prevent bleeding out 5. regulation: control different functions such as regulating blood glucose levels 6. transport: function as carriers of molecules or ions across membranes or between cells 7. storage: serve as a reservoir of essential nutrients 8. stress response: the capacity of living organisms to survive a variety of stresses is mediated by certain proteins; excessively high temperatures and other stresses such as the synthesis of a class of proteins called heat shock proteins (hsps) that promote the correct refolding of damaged proteins if such proteins are severely damaged, hsps degrades it multifuntion proteins (moonlighting proteins): examples are GAPD (catalyzes the oxidation of glyceraldehyde3phosphate; has roles in DNA replication and repair) and crystallin crystallin must prevent the scattering of visible light; have to appear genetically “recruited” from metabolic enzymes, but still retained old functions Protein families: composed of protein molecules that are related by amino acid sequence similarity; have common ancestry superfamilies: proteins that are more distinctly related proteins are often classified based on shape and composition: o fibrous proteins: long rod shaped molecules; insoluble in water and tough; have structural and protective functions EX. keratin o globular proteins: compact, spherical; usually water soluble; dynamic functions o conjugated protein: a simple protein combined with a nonprotein component known as a prosthetic group glycoproteins: contain a carbohydrate component lipoproteins: contain a lipid molecule metalloproteins: contain metal ions phosphoproteins: contain a phosphate group hemoproteins: have heme groups Protein structure: several levels of structural organization of proteins Primary: polypeptides that have similar amino acid sequences and have arisen from the same ancestral gene are homologous; the amino acid residues that are identical in all homologues of a protein are known as invariant and are essential for the protein’s function primary structure, evolution, and molecular diseases: a significant number of primary sequence changes do not affect the function; some substitutions are known as conservative; if it is variable, it has a nonspecific role for the polypeptide’s function conservative and variable sites have been used to track evolutionary relationships (the longer they’ve been diverged, the more differences the polypeptides structure will have) molecular diseases: mutations can be lethal, but not all of them all immediately EX sickle cell the primary structure of a polypeptide is its amino acid sequence, the amino acids are connected by peptide bonds Amino acid residues that are essential for the molecule’s function are referred to as invariant proteins with similar amino acid sequences and functions and a common origin are said to be homologous Secondary: consists of several repeating patterns; most common types are the alphahelix and the betapleated sheets both are stabilized by localized hydrogen bonding between the carbonyl and NH groups in the polypeptide’s backbone; they occur when all the phi (Φ) angles (from the alpha carbonN) and the psi(ψ) angles (from the alpha carbonC) are equal betapleated sheets can be antiparallel (chains are arranged in opposite directions) or parallel (hydrogen bonds in the chains are arranged in the same direction) many globular proteins contain combinations of alphahelix and Bpleated sheet secondary structures; known as supersecondary structure or motifs Tertiary: refers to the unique 3D conformations that globular proteins assume as they fold into their native structures protein folding is unorganized has several features: 1. many polypeptides fold in such a fashion that amino acid residues that are distant from each other in the primary structure come into close proximity 2. globular proteins are compact because of efficient packing as it folds 3. large globular proteins often contain several compact units called domains (structurally independent segments that have specific functions); the core 3D structure of a domain is a fold 4. modular and mosaic proteins contain duplicate or imperfect copies of one or more domains that are linked in series Types of interaction to stabilize tertiary structure: 1. hydrophobic interactions: hydrophobic R groups are brought in close proximity because they are excluded from water 2. Electrostatic interactions: strongest interaction in proteins occurs between ionic groups of opposite charge known as salt bridges are noncovalent bonds which excludes water because of the energy required to remove water molecules 3. Hydrogen bonds: form within the protein’s interior and on its surface; presence of water precludes the formation of hydrogen bonds 4. covalent bonds: created by chemical reactions that alter a polypeptides structure during or after synthesis most prominent in tertiary structure are disulfide bridges 5. Hydration: structured water is an important stabilizing feature of protein structure free energy equation: ΔG =ΔH TΔS o Quaternary: multisubunit proteins in which some or all subunits are identical are oligomers, which are made up of protomers; A large number of oligomeric proteins contain two or four subunits are referred to as dimers and tetramers o several reasons for common occurrence of multisubunit proteins: synthesis of separate subunits may be more efficient than substantially increasing the length of a single polypeptide chain, in supramolecular complexes such as collagen fibers, replacement of smaller worn out or damaged components can be managed more effectively; the complex interactions of multiple subunits help regulate a protein’s biological function o allostery the control of protein function through ligand bind o allosteric transitions ligandinduced conformational changes in a protein o effector/modulator ligands that trigger the proteins o unstructured protein: some proteins are completely or partially unstructured IUPs: intrinsically unstructured proteins natively unfolded proteins: complete lack of ordered structure many involved in the regulation of signal transduction, transcription, translation, and cell proliferation Loss of protein structure: denaturation is the disruption of the structure which may include protein unfolding but doesn’t have to; Christian Anfinsen dealt with reversible denaturation of Bovine pancreatic ribonuclease that is denatured by βmercaptoethanol and 8M urea; during this process, ribonuclease (composed of a single polypeptide and 4 disulfide bridges) completely unfolds and loses all biological activity; can be reversed by removing the denaturing agents with dialysis Denaturing conditions include: 1. strong acids or bases: changes in pH alters hydrogen bonding and salt bridge patterns; as it approaches its isoelectric point, it becomes less soluble and may precipitate from solution 2. organic solvents: watersoluble organic solvents interfere with hydrophobic interactions; so do nonpolar solvents 3. detergents: disrupt hydrophobic interactions and proteins unfold into extended polypeptide chains; they are amphipathic because they have hydrophobic and hydrophilic components 4. reducing agents: in the presence of urea (or other reagents) reducing agents convert disulfide bridges to sulfhydryl groups; disrupts hydrogen and hydrophobic interactions 5. salt concentration: water molecules that interact with the proteins ionizable groups get distracted by/attracted to salt when concentration is high; when its high enough, solvation sphere is removed and protein molecules collect together and precipitate called salting out 6. heavy metal ions: may disrupt salt bridges, protein structure and function; EX. Anemia 7. temperature changes: temperature increases, increasing molecular vibration; after time, the weaker bonds break and cause the protein to unfold 8. mechanical stress: stirring and grinding can disrupt the forces holding the protein together biochemists distinguish four levels of the structural organization of proteins in primary structure, the amino acid residues are connected by peptide bonds the secondary structure of polypeptides is stabilized by hydrogen bonds. Prominent examples are alpha helices and beta pleated sheets tertiary structure is the unique 3D conformation that a protein assumes because of the interactions between amino acid side chains; several types of interaction stabilize tertiary structures proteins that consist of several separate polypeptide subunits exhibit quaternary structure both noncovalent and covalent bonds hold the subunits together; some proteins are partially or completely unstructured The Folding Problem: due to Anfinsens’s experiment, scientist suggested that they could predict the 3D structure of any protein if the physical and chemical properties of the amino acids and the forces that drive the folding process were understood; Sitedirected mutagenesis (a recombinant DNA technique in which specific sequence changes can be introduced into a predetermined position in cloned genes) showed that depending on where the hydrophobic interaction occurred and between which amino acids could cause either no effects to the structure (amino acids on the surface) or serious structural changes (core amino acids) o problems with the model: 1. time constraints 2. complexity recent advances have been made in protein folding research using imaginative combinations of technologies: Circular Dichroism is a type of spectroscopy in which the relationship between motion and structure is probed with electromagnetic radiation, and NMR is an imaging technique that measures the absorption of electromagnetic radiation by atomic nuclei in the presence of a strong magnetic field; these two techniques have shown that proteins do not have one pathway of folding, and depending on the size of the polypeptide it could have intermediates as well; molten globule is a partially organized globular state of a folding polypeptide that resembles the molecule’s native state and within the interior of molten globules the tertiary interaction have not yet stabilized; molecular chaperones: aid living cells during their folding process Molecular chaperones: assist unfolded proteins in two ways: first proteins must be protected during inappropriate proteinprotein interactions; second proteins must fold rapidly and precisely into their correct conformations; there are two major classes of chaperones: 1. Hsp70s: family that bind to and stabilize proteins during the early stages of proteins 2. Hsp60s: once an unfolded polypeptide has been released by hsp70, they are sent to these chaperones that mediate protein folding all the information required for each newly synthesized polypeptide to fold into its new biologically active conformation is encoded in the molecules primary sequence some relatively simple polypeptides fold spontaneously into their native conformations other larger molecules require the assistance of proteins called molecular chaperones to ensure correct folding Fibrous proteins: typically contain high proportions of regular secondary structures such as alpha helices and beta pleated sheet; have structural rather than dynamic roles o Collagen: composed of three lefthanded polypeptide helices that are twisted around each other to form a right handed triple helix; type I collagen molecules (in teeth, bone, skin, and tendons) make up 90% of the collagen found in the human body; consists of large numbers of repeating triplets with the sequence GlyXY normally with x and y being proline and hydroxyproline o Globular proteins: functions involve the precise bind of small ligands or large macromolecules such as nucleic acids or other proteins; well researched examples are myoglobin and hemoglobin myoglobin: found in high concentrations in skeletal and cardiac muscle; gives tissue their red color if they have extremely high concentrations then the muscles will appear brown; composed of globin, which is a single polypeptide chain that has 8 segments of alpha helix; have heme binding unit hemoglobin: spherical molecule found in red blood cells and transport oxygen from the lungs to the tissues of the body; there are different forms of hemoglobin molecules but each has two alphachains and two betachains amino acid sequences of myoglobin and hemoglobin are very different although their 3D configurations are very similar cooperative binding: as the first O2 binds to hemoglobin the binding of another hemoglobin to the same molecule is enhanced; results from changes in hemoglobin’s 3D structure that are initiated when the first O2 binds hemoglobin’s dissociation curve has a sigmoidal shape and myoglobin’s dissociation curve is hyperbolic Bohr effect: the dissociation of oxygen from hemoglobin is enhanced if pH increases; causes oxygen to be delivered to cells in proportion to their needs Globular protein function usually involves bind to small ligands or to other macromolecules the oxygenbinding properties of myoglobin and hemoglobin are determined in part by the number of subunits they contain