1. Amino Acids, Proteins

2. Proteins perform many different functions in the body.
Functions of Proteins

3. Examples of Amino AcidsH
+ │
H3N—C—COO− │
H glycine
CH3 │
H alanine

4. Types of Amino Acids Amino acids are classified as
nonpolar (hydrophobic) with hydrocarbon side chains.
polar (hydrophilic) with polar or ionic side chains.
acidic (hydrophilic) with acidic side chains.
basic (hydrophilic) with –NH2 side chains.
Nonpolar Polar
Acidic
Basic

5. Fischer Projections of Amino Acids
are chiral except glycine.
have Fischer projections that are stereoisomers.
that are L are the only amino acids used in proteins.
L-Alanine D-Alanine L-Cysteine D-Cysteine C H 2 S N O 3

6. Zwitter ions A zwitterion has charged −NH3+ and COO– groups.
forms when both the –NH2 and the –COOH groups in an amino acid ionize in water.
has equal + and – charges at the isoelectric point (pI).
O O
║ ║
NH2—CH2—C—OH H3N—CH2—C—O–
glycine zwitterion of glycine

7. Amino Acids as Acids In solutions more basic than the pI,
the —NH3+ in the amino acid donates a proton.
OH–
H3N—CH2—COO– H2N—CH2—COO–
zwitterion Negative ion
at pI pH > pI
Charge: 0 Charge: 1-

8. Amino Acids as Bases In solution more acidic than the pI,
the COO- in the amino acid accepts a proton. H
H3N—CH2—COO– H3N—CH2—COOH
zwitterion Positive ion
at pI pH< pI
Charge: 0 Charge: 1+

9. pH and ionization H+ OH– + + H3N–CH2–COOH H3N–CH2–COO– H2N–CH2–COO–
H3N–CH2–COOH H3N–CH2–COO– H2N–CH2–COO–
positive ion zwitterion negative ion
low pH pI high pH

10. Separation of Amino Acids
When an electric current is used to separate a mixture of amino acids
the positively charged amino acids move towards the negative electrode.
the negatively charged amino acids move toward the positive electrode.
an amino acid at its pI does not migrate.
the amino acids are identified as separate bands on the filter paper or thin layer plate.

11. Separation of Amino Acids
With an electric current, a mixture of lysine, aspartate, and valine are separated.

12. Learning Check + | | H3N—CH—COOH H2N—CH—COO– (1) (2)
CH CH3
+ | |
H3N—CH—COOH H2N—CH—COO–
(1) (2)
Which structure represents:
A. Alanine at a pH above its pI?
B. Alanine at a pH below its pI?

13. Solution + | | H3N—CH—COOH H2N—CH—COO– (1) (2)
CH CH3
+ | |
H3N—CH—COOH H2N—CH—COO–
(1) (2)
Which structure represents:
A. Alanine at a pH above its pI? (2)
B. Alanine at a pH below its pI? (1)

14. Amino Acids

15. Alanine Alanine has a methyl group for the “R”
Has 3 carbons (glycine had just 2)
NOT essential

16. Serine
NOT essential

17. Threonine
Essential

18. Valine Essential Valine is a branched chain amino acid.
An "ideal protein" would contain 70% as much isoleucine as lysine.
A rare, inherited metabolic disease in which there is a failure of oxidative decarboxylation (breakdown) of valine, leucine and isoleucine results in maple syrup urine disease, named because of a characteristic odor of the urine.

19. Leucine
Leucine is abundant in corn protein and many other grain proteins, and is not therefore a concern to supplement. Only the L isomer has biological value. "Ideal protein" would contain 100% as much leucine as lysine.

20. Isoleucine
Isoleucine is a branched chain amino acid. An "ideal protein" would contain one-half as much isoleucine as lysine.
A rare, inherited metabolic disease in which there is a failure of oxidative decarboxylation (breakdown) of valine, leucine and isoleucine results in maple syrup urine disease, named because of a characteristic odor of the urine.

21. Phenylalanine
Phenylalanine is not tolerated by people with inborn errors of PHE metabolism so they must avoid it. Babies are given a PKU (phenylketonuria) test at birth to detect phenylalanine metabolites in the urine, so they can be treated in order to avoid a dangerous condition.
PHE is combined with ASP (aspartic acid, another amino acid) in the common non-nutritive sweetner aspertame. Equal is another trade name.

22. Methionine Methionine is a sulfur-containing amino acid.
A portion of the methionine requirement can be met from cystine, another sulfur-containing amino acid.
An "ideal protein" would contain one- half as much methionine + cystine as lysine.
Unlike most amino acids, both the d and l isomers of methionine are biologically active. Consequently, it has been synthesized and can be economically purchased for adding to diets.
Methionine participates in acting as a methyl donor, and is involved in the synthesis of many important compounds in the body, including epinephrine and choline.

23. Cysteine
Cystine

24. Tryptophan
Tryptophan is the second most limiting amino acid in corn diets for pigs.
Tryptophan can be used to from the Vitamin niacin but since feeds are usually low in tryptophan and the synthesis is insufficient to meet daily needs, it is not relied upon to meet the animal's need.
Useful in aiding sleep, tryptophan supplements have been banned by FDA as dangerous.
Skatole and indole can be formed from its breakdown in the large intestine by bacteria, producing foul odors.

25. Arginine
Arginine is a precursor of urea in the body. Urea is the form in which nitrogen is removed from mammals, so normal breakdown of protein and deamination of amino acids will result in urea formation, requiring arginine.

26. Histidine
Histidine contains an imidazole group. It is a precursor of histamine. An "ideal protein" would contain 33% as much histidine as lysine.

27. Lysine
Lysine is the most limiting amino acid in corn and most grains for swine and poultry.
Diets balanced for lysine will contain most of the other amino acids in excess of requirement if natural sources of good quality proteins are used.
It can be supplemented in the synthetic form and this is often economical, replacing a portion of the protein needed.
Only the L isomer is biologically active.

28. Essential Amino Acids Clue to remember: PVT TIM HALL
Phenylalanine
Valine
Threonine
Tryptophan
Isoleucine
Methionine
Histidine
Arginine
Lysine
Leucine

29. Amino Acid Metabolism 1. General Features
Nitrogen Balance & Metabolic Pools
2. Degradation
Transamination & Glutamate Dehydrogenases
3. Urea Cycle
4. Sulfur-containing amino acids
5. Creatine & Creatinine

33. Formation of Peptides

34. The Peptide Bond A peptide bond is an amide bond.
forms between the carboxyl group of one amino acid and the amino group of the next amino acid.

35. Formation of a Dipeptide

36. Learning Check
Write the dipeptide Ser-Thr.

37. Solution
Write the dipeptide Ser-Thr.

38. Naming Dipeptides A dipeptide
is named from the free amine (NH3+) using a -yl ending for the name.
names the last amino acid with the free carboxyl group (COO-) by its amino acid name.

39. Learning Check
Write the three-letter abbreviations and names of the tripeptides that could form from two glycine and one alanine.

40. Solution Glycylglycylalanine Gly-Gly-Ala
Write the names and three-letter abbreviations of the tripeptides that could form from two glycine and one alanine.
Glycylglycylalanine Gly-Gly-Ala
Glycylalanylglycine Gly-Ala-Gly
Alanylglycylglycine Ala-Gly-Gly

41. Learning Check
What are the possible tripeptides formed from one each of leucine, glycine, and alanine?

42. Solution Leu-Gly-Ala Leu-Ala-Gly Ala-Leu-Gly Ala-Gly-Leu Gly-Ala-Leu
Tripeptides possible from one each of leucine, glycine, and alanine:
Leu-Gly-Ala
Leu-Ala-Gly
Ala-Leu-Gly
Ala-Gly-Leu
Gly-Ala-Leu
Gly-Leu-Ala

43. Learning Check Write the three-letter abbreviation and name for the
following tetrapeptide.
CH3 │
CH S
│ │
CH–CH3 SH CH2
│ │ │
CH3 O H CH O H CH2O H CH2 O
+ │ ║ │ │ ║ │ │ ║ │ │ ║
H3N–CH–C–N–CH–C–N–CH–C–N–CH–CO–

44. Solution │ CH3 S │ │ CH–CH3 SH CH2 │ │ │ CH3 O H CH O H CH2O H CH2 O
Ala-Leu-Cys-Met Alanylleucylcysteinylmethionine
CH3 │
CH S
│ │
CH–CH3 SH CH2
│ │ │
CH3 O H CH O H CH2O H CH2 O
+ │ ║ │ │ ║ │ │ ║ │ │ ║
H3N–CH–C–N–CH–C–N–CH–C–N–CH–CO–
Ala
Leu
Cys
Met

45. Qualitative Tests for Amino Acids
There are a number of qualitative tests to detect the presence of amino acids
These are largely dependent on the nature of R-group.

46. Exp. 1 Ninhydrin Reaction
A color reaction given by amino acids and peptides on heating with the chemical ninhydrin.
The technique is widely used for the detection and quantitation (measurement) of amino acids and peptides.
Ninhydrin is a powerful oxidizing agent which reacts with all amino acids between pH 4-8 to produce a purple colored-compound.
The reaction is also given by primary amines and ammonia but without the liberation of Co2
The amino acids proline and hydroxyproline also reacts but produce a yellow color.

47. Exp. 1 Ninhydrin Reaction
■ Method:
1 ml AA + 1 ml NH
heat in boiling WB for 5min.
Purple color.

48. Exp. 1 Ninhydrin Reaction
■ Method:
α-amino acid + 2 ninhydrin --->
CO2 + aldehyde + final complex (purple) + 3H2O
In summary, ninhydrin, which is originally yellow, reacts with amino acid and turns deep purple. It is this purple color that is detected in this method.

49. Exp. 2 Xanthoproteic Reaction
This reaction involves the nitration of benzene nucleus in alkaline medium. As a result AAs that contain aromatic nucleus undergo this reaction.
Aromatic AAs form yellow nitro derivative on heating with conc. nitric acid, the salts of these derivatives are orange.
Phenylalanine
Tryptophan
Tyrosine

50. Exp. 2 Xanthoproteic Reaction
■ Method:
1 ml AA + 1 ml conc. HNO3
heat the mixture in WB for 30s
cool
add drop-wise 40% NaOH to render the solution alkaline
Yellow to orange color.

51. Exp. 2 Xanthoproteic Reaction
Tryptophan
Tyrosine
(a)Nitrated tyrosine and tryptophan (b)

52. Exp. 3 Millon Reaction
This reaction is used to detect the presence of phenol (hydroxybenzene) which reacts with Millon's reagent to form red complexes.
The only phenolic AA is tyrosine.
Tyrosine

53. Exp. 3 Millon Reaction 1 ml AA + 5 drops of Millon reagent
■ Method:
1 ml AA + 5 drops of Millon reagent
heat the mixture in BWB for 10min
cool too room temp
add 5 drops of NaNO2
Brick red color.

54. Exp. 4 Hopkin-cole Reaction
This reaction is used to detect the presence of indol group
The indol group of tryptophan reacts with glyoxalic acid in the presence of conc. H2SO4 to give purple color.
Tryptophan

55. Exp. 4 Hopkin-cole Reaction
■ Method:
1 ml AA + 1 ml Hopkin-cole reagent
mix well
Carefully pour conc. H2SO4 down the side of the tube so as to form two layers
Purple ring at the interface.

56. Exp Sulfur Reaction
This reaction is specific to detect the presence of sulfur.
The sulfur of cystein and cystine is converted to inorganic sulfide with conc. NaOH. Lead acetate is added and a ppt of black lead sulfide indicates a +ve reaction.
Cystein

57. Exp Sulfur Reaction
2 ml AA + 1 ml 40% NaOH drops of lead acetate solution
heat the mixture in WB for 3min
Cool
observe any change
Black ppt.

58. Exp. 6 Sakaguchi Reaction
This reaction is used to detect the presence of guanidine group.
The only AA that contains guanidine group is arginine which reacts with α-naphthol and an oxidizing agent such as bromide water to give a red color.
Arginine

59. Exp. 6 Sakaguchi Reaction
2 ml AA + 1 ml 2M NaOH + 1 ml ethanolic 0.02% α-naphthol
mix well
cool in ice
add 1 ml of alkaline hypochlorite solution
Red color