Download document 8294813

Amino Acids, Pep.des, and Proteins Amino acids are the fundamental building blocks of proteins To see how amino acids are incorporated into proteins and the structures of proteins Proteins – Amides from Amino Acids • Chains with fewer than 50 units are called pep.des • Protein: large chains that have structural or cataly.c func.ons in biology Essen.al Amino Acids • All 20 of the amino acids are necessary for protein synthesis • Humans can synthesize only 10 of the 20 • The other 10 must be obtained from food (essen.al amino acids such as: his.dine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine) Types of side chains • Neutral: FiKeen of the twenty have neutral side chains (non-­‐polar or hydrophobic) • Aspar.c acid and Glutamic have a second COOH and are acidic • Lysine, Arganine, His.dine have addi.onal basic amino groups side chains (the N in tryptophan is a very weak base) • Cysteine, Serine, Tyrosine (OH and SH) are weak acids that are good nucleophiles, also known as polar side chains or hydrophilic amino acids The Structure of an Amino Acid An amino acid can never exist as an uncharged compound. D-­‐Sugars and L-­‐Amino Acids Proteins are derived exclusively from L-­‐amino acids His.dine Some amino acids have ionizable hydrogens on their side chains The pI of Alanine The isoelectric point (pI) of an amino acid is the pH at which it has no net charge. Titra.on Curves of Amino Acids • If pKa values for an amino acid are known the frac.ons of each protona.on state can be calculated (Henderson-­‐Hasselbach Equa.on) • pH = pKa – log [A-­‐]/[HA] • This permits a .tra.on curve to be calculated or pKa to be determined from a .tra.on curve pI Depends on Side Chain • The 15 amino acids with thiol, hydroxyl groups or pure hydrocarbon side chains have pI = 5.0 to 6.5 (average of the pKa’s) • Acidic side chains and a lower pI • Basic side chains and higher pI Synthesizing an Amino Acid Using an HVZ Reac.on Reduc.ve Amina.on of α-­‐Keto Acids • Reac.on of an α-­‐keto acid with NH3 and a reducing agent produces an α-­‐amino acid The Amidomalonate Synthesis • Based on malonic ester synthesis. Convert diethyl acetamidomalonate into enolate ion with base, followed by alkyla.on with a primary alkyl halide Hydrolysis of the amide protec.ng group and the esters and decarboxyla.on yields an α-­‐amino acid Synthesizing an Amino Acid Using the N-­‐Phthalimidomalonic Ester Synthesis Synthesizing an Amino Acid Using a Strecker Synthesis Pep.des and Proteins • Proteins and pep.des are amino acid polymers in which the individual amino acid units, called residues, are linked together by amide bonds, or pep.de bonds • An amino group from one residue forms an amide bond with the carboxyl of a second residue Pep.de Linkages • Two dipep.des can result from reac.on between A and S, depending on which COOH reacts with which NH2 we get AS or SA • The long, repe..ve sequence of ⎯N⎯CH⎯CO⎯ atoms that make up a con.nuous chain is called the protein’s backbone • Pep.des are always wricen with the N-­‐terminal amino acid (the one with the free ⎯NH2 group) on the leK and the C-­‐terminal amino acid (the one with the free ⎯CO2H group) on the right • Alanylserine is abbreviated Ala-­‐Ser (or A-­‐S), and serylalanine is abbreviated Ser-­‐Ala (or S-­‐A) Amino Acid Analysis of Pep.des • The sequence of amino acids in a pure protein is specified gene.cally • If a protein is isolated it can be analyzed for its sequence • The composi.on of amino acids can be obtained by automated chromatography and quan.ta.ve measurement of eluted materials using a reac.on with ninhydrin that produces an intense purple color Pep.de Sequencing: The Edman Degrada.on • The Edman degrada.on cleaves amino acids one at a .me from the N-­‐terminus and forms a detectable, separable deriva.ve for each amino acid Pep.de Synthesis • Pep.de synthesis requires that different amide bonds must be formed in a desired sequence • The growing chain is protected at the carboxyl terminal and added amino acids are N-­‐protected • AKer pep.de bond forma.on, N-­‐protec.on is removed Carboxyl Protec.ng Groups • Usually converted into methyl or benzyl esters • Removed by mild hydrolysis with aqueous NaOH • Benzyl esters are cleaved by cataly.c hydrogenolysis of the weak benzylic C–O bond Amino Group Protec.on • An amide that is less stable than the protein amide is formed and then removed • The tert-­‐butoxycarbonyl amide (BOC) protec.ng group is introduced with di-­‐tert-­‐butyl dicarbonate • Removed by brief treatment with trifluoroace.c acid Pep.de Coupling • Amides are formed by trea.ng a mixture of an acid and amine with dicyclohexylcarbodiimide (DCC) Overall Steps in Pep.de Synthesis Automated Pep.de Synthesis: The Merrifield Solid-­‐Phase Technique • Pep.des are connected to beads of polystyrene, reacted, cycled and cleaved at the end Protein Structure • The primary structure of a protein is simply the amino acid sequence. • The secondary structure of a protein describes how segments of the pep.de backbone orient into a regular pacern. • The ter2ary structure describes how the en.re protein molecule coils into an overall three-­‐
dimensional shape. • The quaternary structure describes how different protein molecules come together to yield large aggregate structures α-­‐Helix • α-­‐Helix stabilized by H-­‐bonds between amide N–H groups and C=O groups four residues away β-­‐Pleated Sheet • β-­‐pleated sheet secondary structure is exhibited by polypep.de chains lined up in a parallel arrangement, and held together by hydrogen bonds between chains Denatura.on of Proteins • The ter.ary structure of a globular protein is the result of many intramolecular acrac.ons that can be disrupted by a change of the environment, causing the protein to become denatured • Solubility is dras.cally decreased as in hea.ng egg white, where the albumins unfold and coagulate • Enzymes also lose all cataly.c ac.vity when denatured Enzymes and Coenzymes • An enzyme is a protein that acts as a catalyst for a biological reac.on. • Most enzymes are specific for substrates while enzymes involved in diges.on, such as papain acack many substrates Types of Enzymes by Func.on • Enzymes are usually grouped according to the kind of reac.on they catalyze, not by their structures How Do Enzymes Work? Citrate Synthase • Citrate synthase catalyzes a mixed Claisen condensa.on of acetyl CoA and oxaloacetate to give citrate • Normally Claisen condensa.ons require a strong base in an alcohol solvent but citrate synthetase operates in neutral solu.on