Amino Acids PDF

Bromole ules andCatalysis 1.1 Amino acids and Proteins (buildine Amino acids are compounds containing carbon, hydrogen, oxygen and nitrogen and serve as monomers block) of proteins. As the name implie...

Bromole ules andCatalysis 1.1 Amino acids and Proteins (buildine Amino acids are compounds containing carbon, hydrogen, oxygen and nitrogen and serve as monomers block) of proteins. As the name implies, these compounds contain both an amino group and a carboxyl group. In an a-amino acid, the amino and carboxyl groups are attached to the same carbon atom, which is called the a-carbon The various q-amino acids differ with respect to the side chain (R group) attached to their a-carbon. a-carboxyl group COO H,NC,H a-amino group R Side chain Figure 1.1 General structure of an amino acid. The Rgroup attached to the a-carbon is different in each amino acid. In the simplest case, the R group is a hydrogen atom and amino acid is glycine. With the exception of proline, all a-amino acids have the same general structure. Proline is unique among the a-amino acids because it is not a primary amino acid, but rather is a secondary amino acid, sometimes called an imino acid. Unlike the primary amino acids which contain a primary amino group (-NH,), proline contains a secondary amino group (-NH-). COO COO CO0 H,N CH H,N Cu H HN CoH H = R pCH, CH, Glycine CH, YCH, Proline CH, R CH, NH, Lysine Figure 1.2 Structure of amino acids (glycine and lysine) and imino acid (proline). In u-amino acids, both the amino group and the carboxyl group are attached to the same many naturally OCCurring amino acids not found in a-carbon atom. o protein, have structures that differ from the a-amino these compounds the amino group is attached to a carbon atom other than the a-carbon atom and they are called B, Y, ð, or E-amino acids depending upon the location of the C-atom to which amino group is attached. Amino acids can act as acids and bases Amino acids contain both an amino (-NH,) and acarboxyl (-COOH) group. At physiological pH (~7..2), the u-amino groups are protonated (-NH;) and the u-carboxyl groups are deprotonated (-CO0). Thus, in solution at physiological pH, amino acids are present largely in their zwitterionic ('double ion') form. The ionization state of an amino acid varies with pH. At a very low pH (i.e. (-cOOH)and uncharged whereas u-amino group is strongly acidic protonated solution), (-NH;) the a-carboxyl and positively charged. Thus, group is protonated the overall charge on the molecule is positive. Similarly, at very high pH, the u-carboxyl group is deprotonated (-CO0) and negatively  Biomolec ules and Catalysis charged whereas a-amino group is deprotonated (-NH,) and uncharged. Thus, the overall charge on the molecu is calle is negative. Amino acids are amphiprotic molecules. A molecule that can both accept and donate protons an amphiprotic species. Since the carboxyl group is able to lose a proton and the amino group is able to accept proton, an amino acid can act both as an acid and a base. A substance that can act as either an acid ora base called an amphoteric substance. Thus, anino acids are also amphoteric in nature. COOH coO Coo H,N Ca H H,N Cu- H H,NC,- H R R 1 R Low pH (=1) Intermediate pH (=7) High pH (= 14) Figure 1.3 The acid-base behaviour of an amino acid in solution. The ionization state of amino acids is altered by a change in pH. At low pH, the positively charged species predominates. As the pH increases, the electrically neutral zwitterion becomes predominant. Azwitterion can act as either an acid (proton donor) or a base (proton acceptor). At higher pH, the negatively charged species predominates. Box 1.1 Stereoisomer Isomers are compounds that have the same molecular formula, meaning they contain the same atoms bonded together in different ways. There are two main types of isomers: constitutional isomers, which have different atomic connectivities, and stereolsomers, which have the same atomic connectivity but a different arrangement of atoms in space. Steroisomers are further classified into two types: enantiomers and diastereomers. Two non-identical molecules Have the same molecutar formula Molecules are isomers Do not have the same connectivity Have the same connectivity Molecules are Molecules are constitutional isomers stereoisomers Have non-superimposable mirror images Have superimposabie mirror images Molecules are Molecules are enantiomers diastereomers Molecules that are non-superimposable miror images are called are said to be chiral molecules; they possess enantiomers. Molecules that can exist as enantiomers the property of chirality. Molecules that are not chiral are said to be achiral molecules -without chirality. Achiral essential difference between chiral and achiral molecules are superimposable on their mirror images. What is molecules? The answer is 'symmetry'. 1f a molecule has a planethe symmetry, a center of symmetry (or points of of one or more asymmetric carbons (or other symmetry), or both, it is not chiral. Many chiral molecules contain atoms). An center or chiral center. A carbon atom to which four asymmetric atom is sometimes referred to as an asymmetric differernt groups are bound is an that contalns only one asymmetric carbon is chiral. For molecules with more than asymmetric carbon. A molecule be chiral or achiral. Moreover, an asymmetric carbon is not a one asymmetric carbon, they may have no asymmetric carbons at all. necessary condition for chirality: some chiral molecutes  BIomolecules and Catalysis 4 The -tt enantiomer nomenclature is based on priority rules Nomenclature of enantiomers: The RS system of carbon. The hiake different substituents bonded to an asymmetric is determined by the atomic number of the four atoms at each asymmetric carbon in a molecule can the atomic number. the higher the priority. The arrangement of steps: be assigned using the following diferent substituents (groups) bound to it. 1. Identify an asymmetric carbon and the four (CIP) rules, Atoms with 2. Assign priorities to the four different substituents according to the Cahn-Ingold-Prelog dictates that the highest higher atomic numbers are given higher priority. The convention applied in this text is denoted as 4. priority is designated as 1, while the lowest priority 3. View the molecule along the bond from the asymmetric carbon to the substituent of lowest priority, 4 Consider the clockwise or counterclockwise order of the remaining substituents. If the priorities of these substituents decrease in the clockwise direction, the asymmetric carbon is said to have the Rconfiguration (R for rectus). If the priorities of these substituents decrease in the counterclockwise direction, the asymmetric carbon is said to have the S configuration (S for sinister). Group of lowest prionty is hidden from view Group of lowest prionty is awavy from observer Clockwise (R) The physIcal and chemcal properties of a pair of enantiomers are identical. However, enantiomers are distinguished by their optical activities because they rotate the plane of polarized light by equal amounts in opposite directions. An important point to note is that the R or S configuration does not indicate whether a molecule will exhibit dextrorotatory or levorotatory optical rotation. Amixture containing equal amounts of two enantiomers is called a racemate or racemic mixture. A racemic mixture doesn't rotate such light at all. |Let's consider a malecule with two asymmetric carbons. Each asymmetric carbon can have either the Ror Sconfiguration. With two possibie configurations at each carbon, four stereoisomers are possible. The 2S, 3S, and 2R,3R isomers form a pair of enantiomers as they are non-super1mposable mirror images. Similarly, the 2S,3R, and 2R,3S isomers also constitute an enant1omeric pair. However, the 2S,3S, and 2s,3R pair, as well as the 2R,3R, and 2R,3S pair, are not enantiomers, ind1cating a different stereochemical relationship. Stereoisomers that are not enantiomers are referred toas diastereoisomers, or more simpiy, diastereomers. Diastereomers are not mirror images. They can be chiral or achiral molecules and can arise when structures have two or more asymmetric carbons. Two diastereoisomers are distinct molecules, exh1biting different physical and chemical properties. 2S,3S 4t Enantiomers ’ 2R,3R OH OH H,C--CH CH-CH,- CH, Diastereomers 1 2 3 45 2S, 3R + Enantiomers + 2R, 3S The two pirs of diastereoisomers are different compounds with different narnes and d1fferent properties, while the rotate enantiomers are the same compound with the same properties, they which polarized light. differing only in the direction in  Biomolecules and Catalysis 5 1.11 Absolute configuration All a-amino acids except glycine are chiral molecules. Achiral amino acid can exist in two configurations that are non-superimposable mirror images of each other. These two configurations are called enantiomers. An enantiomer is identified by its absolute configuration.Different systenms have been developed to specify the absolute configuration of a chiral molecule. The configuration and conformation of molecules have distinct meanings. Aitering the configuration of a molecule always entails breaking covalent bonds. Conversely, changing the conformation of a molecule involves rotating about covalent bonds without breaking them. Conformations of a molecule are easily interconvertible and represent the same molecule. COO COO H,N C H H,N H CH, Glycine (achiral amino acid) Alanine (chiral amino acid) DL System The 'DL system' relates the absolute configuration of the a-carbon of an amino acid to that of the 3-carbon aldose sugar glyceraldehyde. Glyceraldehyde has two absolute configurations. When the hydroxylgroup attached to the chiral carbon is on the left in a Fischer projection, the configuration is 'L; when the hydroxyl group is on the right, the configuration is 'D.This refers only to the absolute configuration of the four substituents (atom or group of atoms) bonded to the chiral carbon, not to optical properties of the molecule. CHO CHO HO - C -H H-C -OH CH,OH CH,OH Mirror L-Glyceraldehyde plane D-Glyceraldehyde Figure 1.4 The absolute configuration of glyceraldehyde. In Fisher projection formulas, if the hydroxyl group is located on the right side of the chiral center, the molecule is D. If the hydroxyB group is on the left side, the molecule is L. For all chiral a-amino acids, with the same configuration as that of L-glyceraldehyde are designated L, and confiqurations related to D-glyceraldehyde are designated D. In L-a-amino acids, the a-amino group on the left whereas in D-u-amino acids, the a-amino group is on the right. COO COO H,N Ca--H H-Ca-NH, CH, CH, Mirror L-Alanine plane D-Alanine Flqure 1.5 The absolute configuration of a-amino acid alan1ne is specified by the Dl system in Fisher projection formnulas. The D and L designations specify the configuration of a single reference carbon, For a-amino acids, the reference carbon 0s the a-carbon. All amino acids except glycine exist in these two different absolute confiqurations. However, ail the amino acids which ribosomically incorporated into proteins exhibit L-confiquration. Therefore, they are all L-a-amino acids. The basis for preference for Lamino acids is not known. D-form of amino acids is not found in ribosomically synthesized proteins, although they exist in some peptide antibiotics and tetrapeptide chain of peptidoglycan cell wal.  Biomolecules and Catahss RS system center, the most useful system to describe For compounds with more than one chiral absolute 'RS System. In addition, with this system, we can define the configuration any reference compound. This approach uses priority rules to specify of a chiral compound inconfiguration is the the atomic number of the four different substituents bonded to an configurations. The asymmetric carbon. The higher priority is based on atbsence of the the higher the priority i.e., atoms of higher atomic number bonded to a chiral center are ranked the atomic number, lower atomic number. above those of let's use the amino acid alanine as an example to understand the prioritization of groupS around a chiral carbon To alanine. the a-carbon is bonded to one nitrogen atom (atomic number 7), two carbon atoms (atomic number 6) and one hydrogen atom (atomic number 1). To assign priorities to these groupS, we consider the atomic numbers. The NH, group is given priority 1 since nitrogen has the highest atomic number. Priorities 2 and 3 are assigned to the COOH and CH; groups, respectively, while the hydrogen atom to the chiral carbon are identical, we look at receives priority 4. When two of the atoms bonded the second, third, and so on, atoms to the COOH group has higher priority (atomic establish priority. In this case. number 8) compared to CH, (atomic number 1). Arrange the molecule So that the H atom 1 NH, points into the paper, indicating it is pointing away from you CH, H COOH Clockwise (R) Arrange the molecule so that the H atom NH, points into the paper, indicat1ng it is pointing away from you COOH H CH, Counterclockwise (S) For the RS system naming, it is essential to position the lowestt priority substituent away from the viewer. If the other three groups are arranged in a clockwise direction of decreasing priority, the chiral center is classified as having the R configuration (where R is from the Latin rectus, meaning 'right'). Conversely, if the three groups decrease in priority in a counterclockwise direction, the chiral center is considered to have the S configuration (where S is from the Latin sinister, meaning 'left'). The absolute configquration of the amino acids at the a-carbon is typically described by the DL System rather than the more modern RS system. According to the RS system, all L-amino acids from proteins have an S absolute configuration, with the exception of L-cysteine. The configuration for L-cysteine is R in the RS system because of the presence of sulfur in the side chain. 1.1.2 Optical activity All amino acids, except for glycine, are chiral molecules and, as a result, exhibit optical activity. An optically acu compound has the ability to rotate the plane of plane-polarized light. In unpolarized light, the oscillates in more than one plane; all perpendicular to the direction of propagation. If the electric field ye electric fleld vector oscillates in single plane perpendicular to the direction of propagation, it is called Iinearly or plane polarlzedllght. Optically active molecules are chiral molecules. For a molecule to be considered chiral, it must lack a plane of plane-polarized symmetry or a center of symmetry, or both. An optically active compound can rotate the plane of  Biomolecules and Catalysis 7 plane of polarized clockwise (to the right) or counterclockwise (to the left). Compounds that rotate the light either counterclockwise are denoted by a plus sign (+) or d. Those rotating light clockwise are termed dextrorotatory, quantitative measure levorotatory, designated by a minus sign (-) or I. The optical rotation of a sample is a termed the extent to which polarimeter. The magnitude of optical rotation indicates of its optical activity, measured using a the sign represents the direction of rotation. the plane of polarized light is rotated, while Analyzer Direction of light propagatlon P-*0-1-- Light source Unpolarized light Plane-polarized light Sample tube containing a chiral compound Rotation of the plane-polarized light The degree of rotation of plane-polarized light depends on several factors: the number of optically active molecules (i.e., concentration)present in the path through which the light beam passes (i.e., path length), temperature, solvent used to dissolve the sample, and wavelength. Typically, optical rotations are measured at 20°C ina solvent such as ethanol or chloroform, using light from a sodium lamp with a wavelength of 589 nm. By convention, the concentration of the sample is expressed in grams per milliter, and the path length is in decimeters. Dividing the observed value of optical rotation, through which the light is rotated, by the path length and concentration yields a value denoted as [a]. This value is specific to the compound in question and is known as the compound's specific rotation. rotation [aj = Observed where, CxI [a]{ = the specific rotation of compound at temperature, T (in °C) and wavelength,. (in nm). If the wavelength of the light used is 589 nm, the symbol 'D' (the D line of a sodium lamp) is used, [a]. C= Concentration (in g/ml), l = Path length (in decimetres) Thus, at a particular temperature and for a given wavelength of light, specific rotation is defined as the observed value of optical rotation when plane-polarized light passes through a sample witha path length of 1decimeter and a sample concentration of 1 gram per milliliter. Because the specific rotation is independent of c and I, it is used as the standard measure of optical activity. Problem A pure sample of the naturally occurring, chiralcompound A(0.0s0 g) is dissolved in water (2.0 mL), and the solution is placed in a 0.5 dm cell. The observed value of the optical rotation of this compound is 0.671°. What is the value of specific rotation [a]? Solution: [a] = Observed CxI rotation where, C= 0.050 2 = 0.025 g/mL 0.6710 = 53.68o 0.025 x 0.5 1.1.3 Standard and non-standard amino acids There are hundreds of different amino acids present in the living organisms; however, only 22 different amino acids partlcipate in protein synthesis, which are incorporated ribosomically into proteins. Such amino acids are called standard (or proteinogenic) amino acids. All standard amino acids are L-a-amino acids. These amino acids are specified by simple three letter. Amino acids that occur naturally in cells but are not incorporated ribosomically  Biomole ules and Catalysis proteins are called nonstandard amino acids. Some nonstandard amino acids are constituents of ribosomically post-translational modification of standard amino acids which are but they are generated by orporatedproteins, nthesized ribosomically into proteins. Examples of some of these amino acids are 4-Hydroxyprooline (a derivative desmosine (a derivative of lysine), derivative of lysine), proline), 5-Hydroxylysine (ay-carboxyglutamate (a derivative of glutamate). N-formyimethionine (a erivative of methionine) and particular, their polit. acids can be classified on the properties of their side chain (or R group), in tandard amino polarity of the side chain varies widely from physiological pH (near pH 7). The rtendency to interact with water at polar and hydrophilic. =on-polar and hydrophobic to highly chain Amino acids with non-polar side alanine, valine. standard amino acids, nine amino acids contain non-polar side chain. These are glycine, Among tryptophan. Proline differs from other members in having leucine, isoleucine, proline, methionine, phenylalanine and Phenylalanine and tryptophan have aromatic its side chain bonded to both the nitrogen and the a-carbon atoms. tryptophan has an indole ring. side chains. The side chain of phenylalanine contains a phenyl ring whereas Amino acids with uncharged polar side chain tyrosine. Six amino acids contain uncharged polar side chain - serine, threonine, cysteine, asparagine, glutamine and Three amino acids, serine, threonine and tyrosine contain hydroxyl groups attached to a side chain. Cysteine Is structurally similar to senne but contains a suifhydryl or thiol group (-SH) in place of the hydroxylgroup. Amino acids with charged polar side chain Some standard amino acids with poiar side chains can be positively or negatively charged at neutral pH. Positively charged side chain Lysine and argin1ne have side chains that contain positively charged groups at neutral pH. Lysine has an amino group whereas arginine contains aguanidinium group.Histidine contains an imidazole aromaticring (a planar five member heterocycic ring). The imidazole ring can be uncharged or positiveiy charged near neutral pH, depenain9 on its local env1ronment. Negatively charged side chain Amino acids aspartate and glutamate contain acidic side chains that contain negatively charged carboxyl groups at neutraf pH. Table 1.1 Classificaton of amino acids based on chemical nature of R group Chemical nature of R group Example Aliphatic Giy, Ala, Val, Leu Aromatic Phe, Ty, Trp Hydroxyl Ser, Thr Carboxylic Asp, Giu Sulphur containing Cys, Met Amino Lys, Arg Amide Asr, Gln The structures of the 20 standard amino acIGs, alorng with the chemical nature of rheir R table 1.2. Properties of selenocysteine (5ec) arnd arouDS, are listed pyrrolysine (Pyi), referred to as the 21 st and 22nd amino respectively, are mentioned in the next section.  Biomolecules and Catalysis 9 Amino acids with non-polar side chain Glycine Alanine Valine Leucine Isofeucine COO COO COO COO COO H,N-C-H H,G-C-H H,N-C-H H,N-C-H H,N-C--H CH, CH CH, H-C-CH, / CH, CH, CH CH,CH, CH, CH, Proline Methionine Phenylalanine Tryptophan COoT coo COO COO H,N-C-H H,GC-H H,NC -H H,N-C-H H, CH, CH, CH, CH2 CH, CH, C S HO CH, Amino acids with uncharged polar side chain at pH 7 Cysteine Asparagine Glutamine Tyrosine Serine Threonine COO cO0 COO COO COO cOo H,N-CH H,N-C-H H,N-C-H H,NC-H H,N-C-H CH, CH. CH, H-C-OH H-C-OH CH, SH CH, H CH, NH, OH 7 Amino acids with charged polar side chain at pH Negatvely charged R groups Positively charged R groups Aspatate Glutamate Arginine Histidine Lysine COO COO COO cOO COO H,N-C-H H,N-C-h H,N-C-H H,N-C CH, CH, CH. CH, CH, CH, CH, CH, COO CH, CH, NH CH, C NH. NH, NH. remains uncharged at ph7Q hist1dne has been shown charged, a significant fraction of Note: Aithough the side chain  Standard amino acids Nonia side chain Polar side chain Uncharged at pH 7 Charged at pH 7 Vahne Senne Threonine Isoevone steine rone Negative charge Asoaragne Positive charge Methionine Glutamate Lysine Glutamine pnenvaanne Aspartate (Most acidic) Histidine Tyrosine Arginine (Most basic) Felenocysteine Sec or U) s the 21 standard amino aod. It has a structunre similar to that of cysteine, but it contains elenium rathe than sulphur. It s incorporated into Tpet codon. VGA (a stop codon). polypeptides during translation, However, it is specified by a Selenoysteine has its Own tRNA containing the anticodon UCA and it is formed modifing a seine that nas been attached to the mate cehycrogenase contain selenocysteine tRNA. Enzymes like glutathione peroxidase and selenocysteine in their catalytic center. emino acid. It is simila to ysine and is present in Pyrrolysine (Pyl or O) is the 220° standard some bacterial proteins. It is coded by UAG COdon. Coo H,NCH CH, CH, coo CH, CH NH CH S C=0 Se Cyste.ne Sesenocystene Pyrrolysine froblem Determine whether the follow1ng statements are true or faise. If faise, explain why? All 20 standard aminoacics found in proteins have at least one asyrnmetric carbon atorn.. An equimolar mixture of D- and L-alan1ne does not rotate the plane of polarized light. 3. Alanine obtained from a proten hydrolysate has the same absoiute configuration as D-glyceraldehyde. Solution: False; gBycine has no asymmetric carbon atorm True. Faise: alan1ne obtained from aprotein hydrolys1s has the sarne absolute configuration as L-glyceraldehyde.  Biomolecules and Catalysis 11 1.1.4 Titration of amino acids Because amino acids contain ionizable groups, the predominant ionic form of these molecules in solution depends on the ph. Titration of an amino acid illustrates the effect of pH on the net charge of amino acid. It involves the gradual removal of protons with increasing pH. Let us understand the titration of non-ionizable Rgroup containing and ionizable Rgroup containing amino acids separately. Standard amino acids with Non-ionizable R-group Gly, Ala, Val, Leu, le, Pro, Met, Phe, Trp, Ser, Thr, Asn and Gin Ionizable R-group Asp, Glu, His, Arg, Tyr, Cys and Lys Amino acid with non-ionizable R-group Amino acid with ionizable R-group Gly, Ala, Val, Leu, lle, Pro, Met, Phe, Trp, Ser, Thr, Asn and Gln Asp, Glu, His, Arg, Tyr, Cys and Lys pk, = pk, pk, pk, cOOH COOH H,N H,N-CH PK, = pk, CH, R pK, = pk, CH, pK, = pk, R COOH Titration of non-ionizable R group containing amino acids Non-ionizable R groups containing amino acids (such as glycine and alanine) have two ionizable groups (a-amino and a-carboxyl group) that can undergo protonation and deprotonation. Let us understand the titration of alanine. During titration with a strong base such as NaOH, alanine loses two protons in a stepwise fashion. At a very low pH (i.e., strongly acidic solution), the predominant ionic species of alanine is the fulBy protonated form in which the a-carboxyl group is uncharged and a-amino group is positively charged. Under this condition, the net charge on the molecuie is +1 (Ala*l). However, an increase in the pH results in the deprotonation of a-carboxy! (-COOH) and a-amino(-NH) groups. In the first stage of the titration, the -CoOH group loses its proton. After deprotonation of a-carboxyl group, the net charge on the molecule becomes zero (Ala). Its -COOH group has a pk, (labeled pk,) of 2.34. 15 pH range: Net charge: +1 CoOH coo coO DÃ, 9, H,N C H,N C-H CH, CH, CH, Ala* Ala Ala - Fully protonated Zwitterion Fully deprotonated Fiqure 1.6 At very tow pt, alanine is fuliy protonated, and Ala s the predominant species present. As the pH is raised, the most acidic proton dissociates, this is the carboxyic acid groton, which has the lower pk, The fractions of Aia and Ala° are equal when pH = pk, = 2.3. As the pH is raised further, the second dissociat1on occurs, This is the protonated -amino group, which has the higher pk, The fractions of Ala" and Ala are equal with pt pa, = 9.7. As the pH is raised further, Ala becomes the onty species present.