AMINO ACIDS Amino acids Amino acids are the compounds which contain both an amino (-NH2) and carboxylic (COOH) groups in the same molecule. The carbon atom next to the carboxylic group is regarded as α carbon atom. In the amino acids obtained from proteins the amino group is attached to this carbon atom. Hence they are called α –amino acids. The general structure of α –amino acids is shown below

Alpha - carbon atom

H2N

H

O

C

C

R

carboxylic group OH

side chain

amino group About 20 amino acids are obtained by the hydrolysis of proteins. They differ only in the nature of the side chain (R) its formula is H2N – CH2 – COOH R represents the side chain. Classification of amino acids Amino acids are broadly classified as a). Aliphatic amino acids Ex. H2NCH2COOH i) Glycine

NH2 ii) Alanine

CH3

iii) Serine

H2N

CH

COOH

CH

COOH

b). Aromatic amino acids i) Tyrosine

NH2

OH

CH2

HO

CH2

CHCOOH

NH2 ii) Phenylalanine

CH2

CHCOOH

c). Heterocyclic amino acids

i)

Proline

H2C

CH2

H2C

CHCOOH NH

ii)

CH2

Tryptophan NH

CHCOOH NH2

Aliphatic amino acids may be further classified into following classes depending upon their nature. i). Neutral amino acids : They have one amino group and one carboxyl group in their structure Ex.

1) Glycine

H2NCH2COOH

2) Alanine

H2NCHCOOH CH3

ii). Acidic amino acids : They possess one amino group and two carboxylic acids groups in their structure. Thus they are acidic in nature. Ex. Aspartic acid

Aspartic acid CHCOOH

H2N

CH2COOH iii). Basic amino acids : They contain two amino groups and one carboxylic group. Thy are basic in nature Ex. Lysine

Lysine H2N

CH (CH2)4

COOH NH2

iv). Hydroxy amino acids : They contain one hydroxyl group also Ex. Serine Serine H2N

CHCOOH CH2

OH

v). Sulphur containing amino acids: They contain a sulphur atom Ex. Cysteine

Cysteine H2N

CHCOOH CH2SH

Methods of synthesis of amino acids Proteins on hydrolysis give mixture of amino acids. Various methods are used to isolate each amino acid from the mixture. However several laboratory methods are also developed to synthesis specific amino acids. 1. From α – halogenated acids α – amino acids may be prepared by the action of ammonia on α – chloro or acids

α – bromo

0

ClCHCOOH

+

R alpha - chloro acid

2 NH3

50 c

H2NCHCOOH R amino acid

+

NH4Cl

2. Streckar synthesis An aldehyde is treated with HCN to from corresponding cyanohydrin. This is treated with conc. Ammonia and the resulting α – amino nitrile is hydrolysed with acid to give corresponding amino acids.

H

H CH3

C

+

O

NH3 CH3

HCN

OH

C

-HO 2

acetaldehyde

CN

H

H hydrolysis

CH3

C

NH2

CH3

C

NH2

CN

COOH Amino nitrile can also be prepared in one step by the action of aqueous solution of KCN and amino nitrite alanine NH4Cl mixture upon aldehydes and ketones. 3. Gabriel synthesis An ester of α – halogenated acid is treated with potassium phthalimide and the resulting product is then hydrolysed to give amino acids.

O C NK

+

- KCl

ClCH2COOC2H5

C O O +

C H NHCH2COOC2H5

H2O H2NCH2COOH glycine

C O

+

COOH COOH phtalic acid

+

C2H5OH ethly alcohol

Properties of amino acids Amino acids are generally colourless crystalline solids, soluble in water but insoluble in ether and other organic solvents. They melt at very high temperature but may decompose in melting. All α – amino acids possess asymmetric carbon atom (ex. cept glycine) and thus they are optically active. Zwitter or dipolar nature of amino acids Zwitter ion or a dipolar ion had both positive and negative charges in the same molecule. Since amino acids contain both acidic group (-COOH) and a basic group (-NH2), in aqueous solution, -COOH group donates a proton and the –NH2 group of the same molecule accepts it. This results in the formation of dipolar ion. In neutral solution, the following equilibrium exists.

+ NH2

NH3 -

H

C

COOH

H

R

C

COO

R

Zwitterion The dipolar ion is charged but electrically neutral. Since amino acids contain amino and carboxyl groups they possess the properties of both a base and an acid. Therefore they are amphoteric in nature. Isoelectric point When an ionized amino acid is placed in electric field. It will actually migrate towards cathode or anode depending upon the pH of the medium. In acidic solutions amino acids exist as cation and migrates towards cathode. +

H3NCHCOO

+

+

+

H

H3N

R

CHCOOH R

Zwitter ion cation In basic medium, the amino acid exists as a negative anion and thus migrates towards anode. +

+

+

H3NCHCOO + OH H3N CHCOO + H2O Each amino acid has definite pH at which it does not migrate towards either cathode or anode but exist as a neutral molecule. This pH is called the isoelectric point of the amino acid R the amino acid has its lowest solubility. It is also observed R that all amino acids do and at this pH, Zwitter ion cation

not have the same isoelectric point. Neutral amino acids have isoelectric point ranging from pH 5.5 to 6.3. isoelectric points for acidic amino acids are at lower pH and range about 2.7 to 3.2. For basic amino acids the isoelectric point lies between 7.6 and 10.8 pH. Peptide bond formation The acidic carboxyl group of an amino acid can react with the basic amino group of another amino acid molecule with the elimination of a water molecule. O

This results in the formation of

bond . This known as a peptide linkage C

N

or peptide bond or amide linkage. H

O

H H2N

H

C

C

OH

+

H HN

R

H2N

- H2O C

COOH

R

H

O

C

C

H

R

N

C

H

R

COOH

peptide linkage When two molecules of amino acids are linked by peptide linkage, a dipeptide is formed. This dipeptide can react with next amino acid molecule giving rise to a tripeptide. When several amino acid molecules are linked by peptide linkages a polypeptide is formed. Polypeptide may be partially represented as follows.

H

O

N

R C

C N

C H R

H

H

This peptide bond is the backbone of the polypeptide chain. The amide of peptide group is flat and the carbonyl and amino group lie in one plane having H (of NH) and O (of CO) trans with respect to each other. X – rays studies indicate that the C – N bond length of peptide – CONH is 1.32 A which is shorter than the usual 1.47 A showing slight double bond character. This double bond nature of C – N bond may be due to resonance and because of this there is restricted rotation about C – N bond giving rise to cis – trans isomerism.

H 00

C

C

C

C

NH

N

C

C +

O

O -

Synthesis of polypeptides Bergmann method (Carbobenzoxy method) Dipeptide can be synthesized by the condensation of amino group of noe amino acid with the carboxylic group of the other through the formation of peptide bond. It can be synthesised as shown below.

H H2N

H

O

C

C

OH

+

H HN

H

COOH

CH3

glycine

H2N

C

-HO 2

alanine

H

O

C

C

H

H N

C

H

CH3

COOH

This is only one possibility of such combination. There is a possibility of getting the mixture of dipeptides (such as AlaGly, AlaAla, Gly – Gly) rather than a single desired peptide. In order to condense the two amino acids in the desired manner, the one first reacting amino group of the amino acid should be protected and then other one should have the carboxyl group protected so that when they react there is only one possibility of such combination. Then proper reagent should be used to remove the protective groups after formation of the dipeptide bond.

A number of protecting agents have been recommended. A good protecting agent is characterized by the fact that it can react with amino or carboxylic group easily and can be removed after the synthesis of the dipeptide. Without affecting the peptide bond. Bergmann and zervas in 1932 used a carbobenzoxy derivatives as protecting agent for amino groups. The following sequences illustrates the synthesis of dipeptide using carbobenzoxy chloride for protecting amino group. 1. The amino group of one amino acid (say Glycine) which is not involved in the formation of peptide bond is protected by first reacting the compound with carbobenzoxy chloride to get carbobenzoxy amino acid. Carbobenzoxy is also known as benzylchloro formate and it is prepared by the action of carbonyl chloride on benzyl alcohol.

C6H5CH2OH

+

COCl2

C6H5CH2OCOCl

+

HCl

carbobenzoxy chloride 2. The reagent is condensed with Amino acid to get carbobenzoxy amino acid. It is then treated with PCL5 or SOCl2 to convert into its corresponding acid chloride.

C6H5CH2OCOCl

+

C6H5CH2OCONHCH2COOH

H2NCH2COOH

carbobenzoxy amino acid

glycine PCl5

C6H5CH2OCONHCH2COCl

C6H5CH2OCONHCH2COOH

3. The product formed is treated with another amino acid (say alamine) to form carbobenzoxy peptide.

C6H5CH2OCONHCH2C OCl

+

- HCl H NHCHCOOH CH3 alanine

C6H5CH2OCONHCH2 CONH CHCOOH CH3

4. The carbobenzoxy group is removed by reduction

H2

Pd

C6H5CH2OCONHCH2CONHCHCOOH CH3

H2NCH2CONHCHCOOH

+

C6H5CH3

+

CO2

CH3 The above steps are repeated to synthesis higher peptides. glycylalanine (dipeptide) If the amino acid contains sulphur, then palladium can not be used for catalytic reduction, since sulphur poisons the catalyst. In such cases, the blocking group carbobenzoxy group, can be removed by means of phosponium iodide or sodium in liquid ammonia.