《有机化学》课程教学资源（教材文献，英文版）CHAPTER 27 AMINO ACIDS, PEPTIDES, AND PROTEINS. NUCLEIC ACIDS

27.4 Synthesis of Amino Acids Electrophoresis is used primarily to analyze mix per gram of protein is added to the aqueous buffer. tures of peptides and proteins, rather than individual sds binds to the protein, causing the protein to un- amino acids, but analogous principles apply. because fold so that it is roughly rod-shaped with the they incorporate different numbers of amino acids CH3(CH)1o CH2- groups of SDs associated with the and because their side chains are different, two pe lipophilic portions of the protein. The negatively tides will have slightly different acid-base properties charged sulfate groups are exposed to the water. the and slightly different net charges at a particular pH DS molecules that they carry ensure that all the pro- Thus, their mobilities in an electric field will be differ- tein molecules are negatively charged and migrate ent, and electrophoresis can be used to separate toward the positive electrode. Furthermore, all the them. The medium used to separate peptides and proteins in the mixture now have similar shapes and proteins is typically a polyacrylamide gel, leading to tend to travel at rates proportional to their chain he term gel electrophoresis for this technique length. Thus, when carried out on a preparative scale, A second factor that governs the rate of migra- SDS gel electrophoresis permits proteins in a mixture tion during electrophoresis is the size (length and to be separated according to their molecular weight. shape of the peptide or protein. Larger molecules On an analytical scale, it is used to estimate the mo- move through the polyacrylamide gel more slowly lecular weight of a protein by comparing its elec than smaller ones In current practice, the experiment trophoretic mobility with that of proteins of known is modified to exploit differences in size more than molecular weight differences in net charge, especially in the SDs gel ater, in Section 27 29, we will see how gel elec the trophoresis of proteins. Approximately 1.5 g of trophoresis is used in nucleic acid chemistry the detergent sodium dodecyl sulfate(SDS, page 745) PROBLEM 27.4 Write structural formulas for the principal species present when he pH of a solution containing lysine is raised from 1 to 9 and again to 13 The acid-base properties of their side chains are one way in which individual amino acids differ. This is important in peptides and proteins, where the properties of the substance depend on its amino acid constituents, especially on the nature of the side chains. It is also important in analyses in which a complex mixture of amino acids is eparated into its components by taking advantage of the differences in their proton donating and proton-accepting abilities. 27. 4 SYNTHESIS OF AMINO ACIDS One of the oldest methods for the synthesis of amino acids dates back to the nineteenth century and is simply a nucleophilic substitution in which ammonia reacts with an c acl CH3,H 2NH3 CHCHCO 2-Bromopropanoic acid Ammonia Alanine(65-70%) Ammonium bromide The a-halo acid is normally prepared by the Hell-Volhard-zelinsky reaction(see Sec- tion19.16) PROBLEM 27.5 Outline the steps in a synthesis of valine from 3-methylbutanoic In the Strecker synthesis an aldehyde is converted to an a-amino acid with one more carbon atom by a two-stage procedure in which an a-amino nitrile is an intermediate Back Forward Main MenuToc Study Guide ToC Student o MHHE Website

PROBLEM 27.4 Write structural formulas for the principal species present when the pH of a solution containing lysine is raised from 1 to 9 and again to 13. The acid–base properties of their side chains are one way in which individual amino acids differ. This is important in peptides and proteins, where the properties of the substance depend on its amino acid constituents, especially on the nature of the side chains. It is also important in analyses in which a complex mixture of amino acids is separated into its components by taking advantage of the differences in their proton￾donating and proton-accepting abilities. 27.4 SYNTHESIS OF AMINO ACIDS One of the oldest methods for the synthesis of amino acids dates back to the nineteenth century and is simply a nucleophilic substitution in which ammonia reacts with an -halo carboxylic acid. The -halo acid is normally prepared by the Hell–Volhard–Zelinsky reaction (see Sec￾tion 19.16). PROBLEM 27.5 Outline the steps in a synthesis of valine from 3-methylbutanoic acid. In the Strecker synthesis an aldehyde is converted to an -amino acid with one more carbon atom by a two-stage procedure in which an -amino nitrile is an intermediate. CH3CHCO2H Br 2-Bromopropanoic acid CH3CHCO2 NH3 Alanine (65–70%) 2NH3 Ammonia NH4Br Ammonium bromide H2O 27.4 Synthesis of Amino Acids 1061 Electrophoresis is used primarily to analyze mix￾tures of peptides and proteins, rather than individual amino acids, but analogous principles apply. Because they incorporate different numbers of amino acids and because their side chains are different, two pep￾tides will have slightly different acid–base properties and slightly different net charges at a particular pH. Thus, their mobilities in an electric field will be differ￾ent, and electrophoresis can be used to separate them. The medium used to separate peptides and proteins is typically a polyacrylamide gel, leading to the term gel electrophoresis for this technique. A second factor that governs the rate of migra￾tion during electrophoresis is the size (length and shape) of the peptide or protein. Larger molecules move through the polyacrylamide gel more slowly than smaller ones. In current practice, the experiment is modified to exploit differences in size more than differences in net charge, especially in the SDS gel electrophoresis of proteins. Approximately 1.5 g of the detergent sodium dodecyl sulfate (SDS, page 745) per gram of protein is added to the aqueous buffer. SDS binds to the protein, causing the protein to un￾fold so that it is roughly rod-shaped with the CH3(CH2)10CH2± groups of SDS associated with the lipophilic portions of the protein. The negatively charged sulfate groups are exposed to the water. The SDS molecules that they carry ensure that all the pro￾tein molecules are negatively charged and migrate toward the positive electrode. Furthermore, all the proteins in the mixture now have similar shapes and tend to travel at rates proportional to their chain length. Thus, when carried out on a preparative scale, SDS gel electrophoresis permits proteins in a mixture to be separated according to their molecular weight. On an analytical scale, it is used to estimate the mo￾lecular weight of a protein by comparing its elec￾trophoretic mobility with that of proteins of known molecular weight. Later, in Section 27.29, we will see how gel elec￾trophoresis is used in nucleic acid chemistry. Back Forward Main Menu TOC Study Guide TOC Student OLC MHHE Website

CHAPTER TWENTY-SEVEN Amino Acids, Peptides, and Proteins. Nucleic Acids The a-amino nitrile is formed by reaction of the aldehyde with ammonia or an ammo- nium salt and a source of cyanide ion. Hydrolysis of the nitrile group to a carboxylic acid function completes the synthesis The synthesis of alanine was described by Adolf Strecker CH3CHC≡N 1. H-O. HCl he CH3CHCO, WUrzburg( Germany)in a p. per published in 1850. NH3 Acetaldehyde 2-Aminopropanenitrile Alanine(52-60%) PROBLEM 27.6 Outline the steps in the preparation of valine by the Strecker synthesis The most widely used method for the laboratory synthesis of a-amino acids is a modification of the malonic ester synthesis(Section 21.7). The key reagent is diethyl acetamidomalonate, a derivative of malonic ester that already has the critical nitrogen substituent in place at the a-carbon atom. The side chain is introduced by alkylating diethyl acetamidomalonate in the same way as diethyl malonate itself is alkylated CH, CNHCH(CO, CH CH3) NaocH cH, CH,CNHC(CO, CH, CH3) CH CH c> CH CNHC(CO, CH CH3) CH2C6H Diethyl Sodium salt of Diethyl donate diethyl acetamidomalonate acetamidobenzylmalonate Hydrolysis removes the acetyl group from nitrogen and converts the two ester functions to carboxyl groups. Decarboxylation gives the desired product CH3 CNHC(CO2 CH- CH3)216- H NC(CO2 H)2 -co> CH2CHCOL CHCH CH,C,H NH (not isolated) Phenylalanine acetamidobenzylmalonate (65%) PROBLEM 27.7 Outline the steps in the synthesis of valine from diethyl acetamidomalonate. The overall yield of valine by this method is reported to be rather low(31%). Can you think of a reason why this synthesis is not very effi cient? Unless a resolution step is included, the a-amino acids prepared by the synthetic methods just described are racemic. Optically active amino acids, when desired, may be obtained by resolving a racemic mixture or by enantioselective synthesis. A synthesis is described as enantioselective if it produces one enantiomer of a chiral compound in an amount greater than its mirror image. Recall from Section 7.9 that optically inactive reactants cannot give optically active products. Enantioselective syntheses of amino acids therefore require an enantiomerically enriched chiral reagent or catalyst at some point in Back Forward Main MenuToc Study Guide ToC Student o MHHE Website

The -amino nitrile is formed by reaction of the aldehyde with ammonia or an ammo￾nium salt and a source of cyanide ion. Hydrolysis of the nitrile group to a carboxylic acid function completes the synthesis. PROBLEM 27.6 Outline the steps in the preparation of valine by the Strecker synthesis. The most widely used method for the laboratory synthesis of -amino acids is a modification of the malonic ester synthesis (Section 21.7). The key reagent is diethyl acetamidomalonate, a derivative of malonic ester that already has the critical nitrogen substituent in place at the -carbon atom. The side chain is introduced by alkylating diethyl acetamidomalonate in the same way as diethyl malonate itself is alkylated. Hydrolysis removes the acetyl group from nitrogen and converts the two ester functions to carboxyl groups. Decarboxylation gives the desired product. PROBLEM 27.7 Outline the steps in the synthesis of valine from diethyl acetamidomalonate. The overall yield of valine by this method is reported to be rather low (31%). Can you think of a reason why this synthesis is not very effi- cient? Unless a resolution step is included, the -amino acids prepared by the synthetic methods just described are racemic. Optically active amino acids, when desired, may be obtained by resolving a racemic mixture or by enantioselective synthesis. A synthesis is described as enantioselective if it produces one enantiomer of a chiral compound in an amount greater than its mirror image. Recall from Section 7.9 that optically inactive reactants cannot give optically active products. Enantioselective syntheses of amino acids therefore require an enantiomerically enriched chiral reagent or catalyst at some point in HBr H2O, heat heat CO2 Phenylalanine (65%) C6H5CH2CHCO2 NH3 Diethyl acetamidobenzylmalonate CH3CNHC(CO2CH2CH3)2 O CH2C6H5 (not isolated) H3NC(CO2H)2 CH2C6H5 CH3CH O Acetaldehyde NH4Cl NaCN 2-Aminopropanenitrile CH3CHC N NH2 Alanine (52–60%) CH3CHCO2 NH3 1. H2O, HCl, heat 2. HO 1062 CHAPTER TWENTY-SEVEN Amino Acids, Peptides, and Proteins. Nucleic Acids CH3CNHCH(CO2CH2CH3)2 O Diethyl acetamidomalonate CH3CNHC(CO2CH2CH3)2 Na O Sodium salt of diethyl acetamidomalonate NaOCH2CH3 CH3CH2OH Diethyl acetamidobenzylmalonate (90%) CH3CNHC(CO2CH2CH3)2 O CH2C6H5 C6H5CH2Cl The synthesis of alanine was described by Adolf Strecker of the University of Würzburg (Germany) in a pa￾per published in 1850. Back Forward Main Menu TOC Study Guide TOC Student OLC MHHE Website

27.6 Some biochemical reactions of amino acids the process. If the chiral reagent or catalyst is a single enantiomer and if the reaction sequence is completely enantioselective, an optically pure amino acid is obtained Chemists have succeeded in preparing a-amino acids by techniques that are more than 95%0 enantioselective Although this is an impressive feat, we must not lose sight of the fact that the reactions that produce amino acids in living systems do so with 100% enan- tioselectivity 27.5 REACTIONS OF AMINO ACIDS no acids undergo reactions characteristic of both their amine and carboxylic acid ional groups. Acylation is a typical reaction of the amino group H,NCH, C0,-+ CH.COCCH,-, CH, CNHCH, CO-H +CH,COH N-Acetylglycine(89-92%) Acetic acid Ester formation is a typical reaction of the carboxyl group. CH3CHCO2 CH3CH,OH CH3CHCOCH,CH3 CI NH Alanine Ethanol Hydrochloride salt of alanine The presence of amino acids can be detected by the formation of a purple color Ninhydrin is used to detect on treatment with ninhydrin. The same compound responsible for the purple color is fingerprints formed from all amino acids in which the a-amino group is primary RCH + H3NCHCO2+HO一 4H,O Ninhydrin “ Ruhemann' s purple”) not normally Proline, in which the a-amino group is secondary, gives an orange compound on reac tion with ninhydrin PROBLEM 27.8 Suggest a reasonable mechanism for the reaction of an a-amino acid with ninhydrin 27.6 SOME BIOCHEMICAL REACTIONS OF AMINO ACIDS The 20 amino acids listed in Table 27. 1 are biosynthesized by a number of different pathways, and we will touch on only a few of them in an introductory way. We will examine the biosynthesis of glutamic acid first, since it illustrates a biochemical process Back Forward Main MenuToc Study Guide ToC Student o MHHE Website

the process. If the chiral reagent or catalyst is a single enantiomer and if the reaction sequence is completely enantioselective, an optically pure amino acid is obtained. Chemists have succeeded in preparing -amino acids by techniques that are more than 95% enantioselective. Although this is an impressive feat, we must not lose sight of the fact that the reactions that produce amino acids in living systems do so with 100% enan￾tioselectivity. 27.5 REACTIONS OF AMINO ACIDS Amino acids undergo reactions characteristic of both their amine and carboxylic acid functional groups. Acylation is a typical reaction of the amino group. Ester formation is a typical reaction of the carboxyl group. The presence of amino acids can be detected by the formation of a purple color on treatment with ninhydrin. The same compound responsible for the purple color is formed from all amino acids in which the -amino group is primary. Proline, in which the -amino group is secondary, gives an orange compound on reac￾tion with ninhydrin. PROBLEM 27.8 Suggest a reasonable mechanism for the reaction of an -amino acid with ninhydrin. 27.6 SOME BIOCHEMICAL REACTIONS OF AMINO ACIDS The 20 amino acids listed in Table 27.1 are biosynthesized by a number of different pathways, and we will touch on only a few of them in an introductory way. We will examine the biosynthesis of glutamic acid first, since it illustrates a biochemical process Ethanol CH3CH2OH Alanine CH3CHCO2 NH3 Hydrochloride salt of alanine ethyl ester (90–95%) CH3CHCOCH2CH3 Cl O NH3 HCl Glycine H3NCH2CO2 Acetic anhydride CH3COCCH3 O O N-Acetylglycine (89–92%) CH3CNHCH2CO2H O Acetic acid CH3CO2H 27.6 Some Biochemical Reactions of Amino Acids 1063 2 O O OH OH Ninhydrin H3NCHCO2 R HO N O O O O Violet dye (“Ruhemann’s purple”) (Formed, but not normally isolated) O RCH CO2 4 H2O Ninhydrin is used to detect fingerprints. Back Forward Main Menu TOC Study Guide TOC Student OLC MHHE Website

CHAPTER TWENTY-SEVEN Amino Acids, Peptides, and Proteins. Nucleic Acids analogous to a reaction we have discussed earlier in the context of amine synthesis, reductive amination(Section 22.11) Glutamic acid is formed in most organisms from ammonia and a-ketoglutaric acid a-Ketoglutaric acid is one of the intermediates in the tricarboxylic acid cycle(also called the Krebs cycle)and arises via metabolic breakdown of food sources-carboh Journal of chemical educa. drates, fats, and prote tion(pp 673-677)contains a review of the Krebs cycle HO, CCH, CH,CCO,H+ NH3 HO, CCH,CH, CHCO, NH Ammonia L-Glutamic acid Ammonia reacts with the ketone carbonyl group to give an imine(C=NH), which is then reduced to the amine function of the a-amino acid both imine formation and reduc tion are enzyme-catalyzed. The reduced form of nicotinamide adenine diphospho cleotide(NADPh) is a coenzyme and acts as a reducing agent. The step in which the imine is reduced is the one in which the stereogenic center is introduced and gives only -glutamic aci L-Glutamic acid is not an essential amino acid. It need not be present in the diet, since animals can biosynthesize it from sources of a-ketoglutaric acid. It is, however, key intermediate in the biosynthesis of other amino acids by a process known as transamination L-Alanine, for example, is formed from pyruvic acid by transamination from L-glutamic acid O CH=CCO2H+ HOCCH, CH,CHCO > CH3CHCO2 t HO2CCH,CH, CCO NH3 Pyruvic acid L-Glutamic acid L-Alanine a-Ketoglutaric acid In transamination an amine group is transferred from L-glutamic acid to pyruvic acid An outline of the mechanism of transamination is presented in Figure 27.4 One amino acid often serves as the biological precursor to another. L-Phenylala nine is classified as an essential amino acid, whereas its p-hydrox xy derivative, L-tyro- sine, is not. This is because animals can convert L-phenylalanine to L-tyrosine by hydrox ylation of the aromatic ring. An arene oxide(Section 24.7)is an intermediat CHCHCO NH NH3 NH Arene oxide intermediate L-Tyrosine bolic pathway, giving phenylpyruvic acl B ary to convert L-phenylalanine to L-tyrosine. Some people lack the enz mes Any L-phenylalanine that they obtain from their diet is diverted along a different meta Back Forward Main MenuToc Study Guide ToC Student o MHHE Website

analogous to a reaction we have discussed earlier in the context of amine synthesis, reductive amination (Section 22.11). Glutamic acid is formed in most organisms from ammonia and -ketoglutaric acid. -Ketoglutaric acid is one of the intermediates in the tricarboxylic acid cycle (also called the Krebs cycle) and arises via metabolic breakdown of food sources—carbohy￾drates, fats, and proteins. Ammonia reacts with the ketone carbonyl group to give an imine (CœNH), which is then reduced to the amine function of the -amino acid. Both imine formation and reduc￾tion are enzyme-catalyzed. The reduced form of nicotinamide adenine diphosphonu￾cleotide (NADPH) is a coenzyme and acts as a reducing agent. The step in which the imine is reduced is the one in which the stereogenic center is introduced and gives only L-glutamic acid. L-Glutamic acid is not an essential amino acid. It need not be present in the diet, since animals can biosynthesize it from sources of -ketoglutaric acid. It is, however, a key intermediate in the biosynthesis of other amino acids by a process known as transamination. L-Alanine, for example, is formed from pyruvic acid by transamination from L-glutamic acid. In transamination an amine group is transferred from L-glutamic acid to pyruvic acid. An outline of the mechanism of transamination is presented in Figure 27.4. One amino acid often serves as the biological precursor to another. L-Phenylala￾nine is classified as an essential amino acid, whereas its p-hydroxy derivative, L-tyro￾sine, is not. This is because animals can convert L-phenylalanine to L-tyrosine by hydrox￾ylation of the aromatic ring. An arene oxide (Section 24.7) is an intermediate. Some people lack the enzymes necessary to convert L-phenylalanine to L-tyrosine. Any L-phenylalanine that they obtain from their diet is diverted along a different meta￾bolic pathway, giving phenylpyruvic acid: enzymes L-Glutamic acid HO2CCH2CH2CHCO2 NH3 CH3CHCO2 NH3 Pyruvic acid L-Alanine CH3CCO2H O -Ketoglutaric acid HO2CCH2CH2CCO2H O enzymes reducing agents L-Glutamic acid HO2CCH2CH2CHCO2 NH3 -Ketoglutaric acid HO2CCH2CH2CCO2H O Ammonia NH3 1064 CHAPTER TWENTY-SEVEN Amino Acids, Peptides, and Proteins. Nucleic Acids The August 1986 issue of the Journal of Chemical Educa￾tion (pp. 673–677) contains a review of the Krebs cycle. CH2CHCO2 NH3 L-Phenylalanine O2 enzyme enzyme O CH2CHCO2 NH3 Arene oxide intermediate HO CH2CHCO2 NH3 L-Tyrosine Back Forward Main Menu TOC Study Guide TOC Student OLC MHHE Website

27.6 Some biochemical reactions of amino acids 1065 Step 1: The amine function of L-glutamate reacts with the ketone carbonyl of pyruvate to form an imine OCCH.CH O CCH.CH CH一NH3+O=C CH-N CH OC CH3 ultimate Step 2: Enzyme-catalyzed proton-transfer steps cause migration of the double bond, converting the imine formed in step I to an isomeric imine O, CCH, CH, H CO OCCH CH H CO, CH CH3 Rearranged imine Step 3: Hydrolysis of the rearranged imine gives L-alanine and a-ketoglutarate O,,CH CO O. CCHCH C=N一CH+HO一 O+HN一CH Oc CH CH3 Water a-Ketoglutarate L-Alanine FIGURE 27. 4 The mechanism of transamination. All the steps are enzyme-catalyzed CHCHCO CH,CCO,H NH3 Phenylalanine Phenylpyruvic acid Phenylpyruvic acid can cause mental retardation in infants who are deficient in the enzymes necessary to convert L-phenylalanine to L-tyrosine. This disorder is called phenylketonuria, or PKU disease. PKU disease can be detected by a simple test rou- tinely administered to newborns. It cannot be cured, but is controlled by restricting the dietary intake of L-phenylalanine. In practice this means avoiding foods such as meat that are rich in L-phenylalanine. Among the biochemical reactions that amino acids undergo is decarboxylation to amines. Decarboxylation of histidine, for example, gives histamine, a powerful vasodila tor normally present in tissue and formed in excessive amounts under conditions of trau- matic shock Back Forward Main MenuToc Study Guide ToC Student o MHHE Website

Phenylpyruvic acid can cause mental retardation in infants who are deficient in the enzymes necessary to convert L-phenylalanine to L-tyrosine. This disorder is called phenylketonuria, or PKU disease. PKU disease can be detected by a simple test rou￾tinely administered to newborns. It cannot be cured, but is controlled by restricting the dietary intake of L-phenylalanine. In practice this means avoiding foods such as meat that are rich in L-phenylalanine. Among the biochemical reactions that amino acids undergo is decarboxylation to amines. Decarboxylation of histidine, for example, gives histamine, a powerful vasodila￾tor normally present in tissue and formed in excessive amounts under conditions of trau￾matic shock. CH2CHCO2 NH3 L-Phenylalanine CH2CCO2H Phenylpyruvic acid enzymes O 27.6 Some Biochemical Reactions of Amino Acids 1065 Step 1: The amine function of L-glutamate reacts with the ketone carbonyl of pyruvate to form an imine. L-Glutamate Step 2: Enzyme-catalyzed proton-transfer steps cause migration of the double bond, converting the imine formed in step 1 to an isomeric imine. Pyruvate Imine Imine from step 1 H H H±acid Step 3: Hydrolysis of the rearranged imine gives L-alanine and -ketoglutarate. Rearranged imine Rearranged imine Water -Ketoglutarate L-Alanine O2CCH2CH2 O2CCH2CH2 CH±NH3 OœC CH±NœC O2C O2CCH2CH2 C±NœC O2C O2C CO2 CH3 CO2 CH3 CO2 CH3 O2CCH2CH2 CœN±C O2C CO2 CH3 ¢± O2CCH2CH2 CœO H3N±CH O2C CO2 CH3 ¢± ¢± O2CCH2CH2 CœN±CH H2O O2C CO2 CH3 base: FIGURE 27.4 The mechanism of transamination. All the steps are enzyme-catalyzed. Back Forward Main Menu TOC Study Guide TOC Student OLC MHHE Website