Created by Dr Elena D. Korir

LECTURE 1. AMINO ACIDS AND PROTEINS

Amino acids constitute the proteins. Most proteins contain 20 L-α-amino amino acids.

Essential Amino Acids:

The human diet must contain adequate quantities of ten essential amino acids, since neither human nor any other higher animal can synthesize these amino acids in amounts adequate to support infant growth or maintain health in adults.

Ile, Leu, Lys, Met, Phe, Thr, Trp and Val

Other less common modified or “unusual” amino acids also fulfill highly specific functions in the proteins in which they occur.

Properties of Amino Acids

1) Ionization of Amino Acids

In solution, two forms of these groups, one charged and one uncharged, exist in protonic equilibrium.

R-COOH is a far stronger acid than is R-NH3+

At the pH of blood plasma or the intracellular space (7.4 and 7.1 respectively), carboxyl groups exist almost entirely as carboxylate ions, R-COO-. At these pH values, most amino groups are predominantly in the protonated form, R-NH3+.

The net charge of an amino acid depends upon the pH, or proton concentration, of the surrounding solution.

2) Reaction of Amino Acids to Water

3)

4) Condensation Reaction

NH3+-CH2-COO- + NH3+-CH-COO- NH3+-CH2-CO-NH-CH-COO-

CH3 CH3

Glycine Alanine Dipeptide

Peptide bond

Peptides

Proteins >50AA

5) Cys-SH + Cys-SH Cys-S-S-Cys + H2

disulphide bridge

CHEMICAL PROPERTIES OF PROTEINS

1 Acid hydrolysis

[H+]

Proteins Amino Acids

Proteins Peptides Amino Acids

E: Pepsin/Trypsin/Chymotrypsin

2 Denaturation

Turbidity test for proteins in urine in clinical biochemistry

Protein + TCA Denatured protein

3 Test for N-terminal Amino Acid

4 Test for C-terminal Amino Acid

H2N-CH(R1)-CO-NH-CH(R2)-CO-NH-CH(R3) COOH

E: Carboxypeptidase (A,B.C.Y)

H2N-CH(R1)-CO-NH-CH(R2)-COOH + H2N-CH(R3)-COOH

Carboxypeptidase A,B and C extracted from pancreas

5 Reaction of Carbohydrates and Proteins

N-glycosidic linkage is through the amide group of Asn.

The O-glycosidic bond is to the hydroxyl of Ser, Thr or hydroxylysine (hLys).

6 Lipoproteins

7 Phosphoproteins

8 Biuret Test

NaOH

Proteins + Cu(OH)2 Proteins-CuO Complex

Blue-purple colour

The colour of the complex is proportional to the number of peptides bonds and concentration of the protein. The Biuret reaction can be used to assess the concentration of proteins because peptide bonds occur with the same frequency per amino acid in the peptide. The intensity of the color, and hence the absorption at 540 nm, is directly proportional to the protein concentration, according to the Beer-Lambert law.

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Dansyl Chloride Reaction

N(CH

3

)

2

SO

2

Cl

+

NH

2

CH

R

1

C

O

NH

-

=

CH

R

2

-

C=O

NH

CH

-

R

3

-HCl

(base)

SO

2

N(CH

3

)

2

R

3

-

CH

NH

C=O

R

2

-

CH

=

-

NH

O

C

R

1

CH

NH

NH

CH

R

1

C

O

NH

-

=

CH

R

2

-

CO

NH

CH

-

R

3

N(CH

3

)

2

SO

2

OH

+

+

COOH

COOH

Tripeptide

Dansyl tripeptide

Free amino acids

2

2

2H

2

O

(acid)

Dansyl amino acid

O

CH

2

OH

OH

+ H

2

N-CH(R

1

)-CO-

-

-

HN-CH(R

1

)-CO-

CH

2

OH

O

protein

N-glycoside (glycoprotein)

O

CH

2

OH

OH

+ H

2

N-CH(R

1

)-CO-

-

-

HN-CH(R

1

)-CO-

CH

2

OH

O

protein

N-glycoside (glycoprotein)

O

CH

2

OH

OH

-HN-CH-CO-

-

-

O-CH

2

-CH-CO-

CH

2

OH

O

O-glycoside (glycoprotein)

CH

2

OH

+

NH

Ser-protein

O

CH

2

OH

OH

-HN-CH-CO-

-

-O-CH

2

-CH-CO-

CH

2

OH

O

O-glycoside (glycoprotein)

CH

2

OH

+

NH

Ser-protein

R-COOH + H

2

N-CH(R

1

)-CO-

-

-

R-CO-HN-CH(R

1

)-CO-

fatty acid

protein

lipoprotein

R-COOH + H

2

N-CH(R

1

)-CO-

-

-

R-CO-HN-CH(R

1

)-CO-

fatty acid

protein

lipoprotein

-

-

NH-CH(CH

2

)-CO-

-

+ H

3

PO

4

OH

O

-

NH-CH(CH

2

)-CO-

-

-

HO-P-OH

O

protein (Ser)

phosphoprotein (Ser-phosphate)

milk

-

-

NH-CH(CH

2

)-CO-

-

+ H

3

PO

4

OH

O

-

NH-CH(CH

2

)-CO-

--

HO-P-OH

O

protein (Ser)

phosphoprotein (Ser-phosphate)

milk

R-CH-NH

2

COOH

R-COOH

R-COO

-

+ H

+

R-NH

3

+

R-NH

2

+ H

+