Amino acids and protein chemistry 1

AMINO ACIDS

AND

PROTEIN

CHEMISTRY

GENERAL STRUCTURE/FORMULA OF AMINO ACIDS

COO- | +H3N- C – H | R

α- Carbon

Amino group

Carboxyl

group Hydrogen

Side chain

1. α-carbon bonded to an amino group, a

carboxyl group, a hydrogen atom and a

side-chain group.

2. The amino group bonded to the α-carbon

makes all AAs for that matter as α-amino

acids.

3. R group – determines the identity of the

AA and the role or biologic property of the

AA in a protein molecule.

4. Carbon atoms of the side chains are

sequentially labeled as β, γ, δ, ε, which

refer to carbons 3, 4, 5, and 6 respectively.

5. α-carbon is a chiral or asymmetric carbon

because there are 4 different groups

attached to it.

6. All AAs except glycine have chiral

carbons; glycine has 2 H atoms attached

to the α-carbon, hence a symmetric

molecule.

7. The amino and carboxyl groups

participate in chemical reactions, hence

constitute the functional groups of amino

acids and are unique to all amino acids.

7. At physiologic ph (7.4), the COOH group is

deprotonated, forming the negatively charged

carboxylate ion ( -CO0-) and the amino group

is protonated (-NH3+).

8. With each AA containing at least one amino

and carboxylic acid group, amino acids are

classified as amphoteric substances and react

with both acids and bases.

http://en.wikipedia.org/wiki/Amphoterism

D & L FORMS (ISOMERS) OF AMINO ACIDS

1. Glyceraldehyde is used as a reference compound for D and L isomers of

amino acids (similar to carbohydrates).

2. In L-glyceraldehyde, the OH- group is on the left side of the molecule;

in D-glyceraldehyde, the OH- group is on the right side.

3. In L-amino acid as L-alanine, the amino group (NH3+) is on the left side

with the carboxyl group at the top of the structure; in D-alanine, the amino

group is on the right side with the carboxyl group at the top of the structure.

4. In proteins, all of the AAs are in the L-isomer; hence by convention, all AAs

are presumed to be in the L configuration unless specifically designated.

CLASSIFICATION OF AMINO ACIDS- based on the

chemical properties of the side chains

Grp I. AAs with Non-polar or Hydrophobic Side Chains

– have the tendency to cluster away from H2O (or H2O-hating)

1. Leucine – have 4-carbon side chain; the purely ketogenic amino acid.

2. Proline

a. Its 3-carbon side chain is bonded to the nitrogen of its α-amino group and to the α-

carbon, creating a cyclic or ring structure; this amino acid contains a secondary

rather than a primary amino group, hence an imino acid or a secondary amine.

b. This substituted α-amino group influences protein folding by forcing a bend in the

polypetide chain.

Imino

group

CLASSIFICATION OF AMINO ACIDS Grp I. AAs with Non-polar Side Chains – also

thought of as “oily” or lipid-like.

Alanine – has a methyl group side chain.

CLASSIFICATION OF AMINO ACIDS Grp I. AAs with Non-polar Side Chains

1. Methionine – side chain contains a sulfur group, similar to cysteine

2. Tryptophan – side chain contains an indole ring; classified as a neutral amino acid.

In Hartnup’s Disease, there is inability of the intestinal epithelial cells to absorb neutral amino acids

like tryptophan excessive amount in the urine impaired synthesis of niacinamide.

3. Phenylalanine – hydrocarbon group is aromatic; i.e., contains a cyclic group similar to a benzene

ring.

CLASSIFICATION OF AMINO ACIDS Grp 2. AAs with Neutral Polar Side Chains (Uncharged) – participate in

hydrogen bonding

1. Glycine – with very small side chain, hence it causes the least hindrance in a

protein (i.e., it does not significantly impinge on the space occupied by other atoms

or chemical groups); the simplest AA.

2. Serine – polar hydroxyl (OH-) group is bonded to aliphatic hydrocarbon groups.

3. Asparagine & Glutamine – with amide groups in their side chains.

CLASSIFICATION OF AMINO ACIDS Grp 2. AAs with Neutral Polar Side Chains (Uncharged)

Threonine – with 2 asymmetric

carbons

Cysteine – polar grp. consists of a –SH

(thiol) grp. which can react with

other cysteine –SH grps. to form

disulfide bridges (-S-S-) or bonds in

proteins.

Tyrosine – hydroxyl grp. is bonded to

an aromatic hydrocarbon grp.

CLASSIFICATION OF AMINO ACIDS Grp 3. AAs with Carboxyl Groups in their Side

Chains (Acidic)

The carboxyl grp. (in addition to the one present in all

AAs, hence dicarboxylic amino acids) makes these

amino acids negatively charged.

CLASSIFICATION OF AMINO ACIDS

Grp 4. AAs with Basic Side Chains – positively charged

1. Histidine – side chain consists of

an imidazole group

2. Arginine – side chain basic group

(guanidino group) is bonded to an

aliphatic hydrocarbon tail; the most

basic amino acid.

3. Lysine – side chain amino group

is attached to an aliphatic hydro-

carbon tail

FORMATION OF THE PEPTIDE BOND

Slide shows the

peptide bond (C-N)

formed between the

carboxyl group of

valine and the amino

group of alanine to

form a dipeptide, read

as valylalanine, not

alaninevaline.

Again, each component

AA in the dipeptide is

called a “residue” or

“moiety”.

PRIMARY STRUCTURE

Refers to the linear sequence of amino

acids in a polypeptide chain

LEU-GLY-THR-VAL-ARG-ASP-HIS

VAL-HIS-ASP-LEU-GLY-ARG-THR

1. Although these 2 peptides have the same number and kinds of

amino acids, they have different sequence of amino acids and

hence have different primary structure.

2. The primary structure determines its 3-dimensional structure,

properties and functions of a protein.

3. This primary structure is determined from the genetic information

encoded in DNA.

SECONDARY STRUCTURE Refers to the hydrogen-bonded arrangement of the backbone of the

polypeptide chain.

1. There are 2 bonds within an AA with

reasonably free rotation:

a. bond bet. the α-carbon and the amino

nitrogen of the residue

b. bond bet. the α-carbon and the

carboxyl carbon of that residue.

2. A peptide chain backbone can be visualized

as a series of playing cards, each card

representing a planar peptide group.

3. The cards are linked at opposite corners by

swivels, representing the bonds about which

there is a considerable rotation.

4. The angles φ (phi) and Ψ (psi), frequently

called Ramachandran angles, are used to

designate rotation around the C-N (α-carbon

and amino nitrogen) and C-C (α-carbon and

carboxyl carbon) bonds respectively.

5. The side chains also play vital role in

determining the 3-dimensional shape of the

protein, but only the backbone is considered

in the secondary structure.

TERTIARY STRUCTURE

OF PROTEINS

1. Refers to the shape of the fully folded

polypeptide chain, hence distant

portions of the secondary structures

are close together.

2. Exemplified by the structure of

MYOGLOBIN

a. Myoglobin is a compact structure

consisting of a single polypeptide

chain (153 AAs) and the prosthetic

group heme-containing Fe.

b. It consists of 8 helices (A to H)

stabilized by hydrogen bonds and

so are the AA side chains.

QUARTERNARY STRUCTURE

OF PROTEINS 1. Refers to the arrangement of 2 or more

polypeptide chains or subunits with

respect to one another to form a

multisubunit molecule.

2. Exemplified by the structure of

HEMOGLOBIN

a. Hemoglobin is a tetramer consisting of 4 polypeptide chains:

*2 α-chains (blue color) – each chain

is 141 residues long

*2 β-chains (green color) – each

chain is 146 residues long

b. The 2 α-chains are identical; the 2 β-

chains are likewise identical.

c. Hemoglobin has therefore this

overall structure: α2β2.

d. Buried in a crevice within each of

the α and β-chains are the prosthetic

groups hemes containing Fe+2.

DENATURATION OF PROTEINS

1. Refers to the unfolding of the protein

hence destruction of its native

conformation, esp. of the secondary

and tertiary structure.

2. This is not accompanied by

hydrolysis of the peptide bonds.

3. Agents that cause denaturation:

a. Heat causes vibrations disruption

of tertiary structure unfolding or

denaturation.

b. Extremes of pH decrease

electrostatic interactions that

maintain native, active form

denaturation.

c. Chemical Agents:

c1. Chaotropic – urea and

guanidinium salts

c2. Detergents – sodium didecyl

sulfate (DDS)

PROTEIN DIGESTION

AND AMINO ACID

ABSORPTION

OVERVIEW OF DIGESTION OF DIETARY PROTEINS

Food

Protein

Polypeptides HCO3

-

Trypsin

Chymotrypsin

Elastase

Carboxypeptidases

A & B

Aminopeptidases

Di- and tripeptides

+ Amino acids

Intestinal epithelial cell

Di- and tri-

peptidases Amino

acids Amino

acids

Oligopeptides

HCl Pepsin

Stomach

Pancreas

Small

intestine

Blood

1. Proteins are too large to be absorbed,

hence they must be first hydrolyzed to

their constituent amino acids.

2. Digestion occurs via the sequential

action of proteolytic enzymes in the

stomach, pancreas & small intestines.

a. In the stomach, dietary proteins are

converted to polypeptides & some

free amino acids by pepsin.

b. In the small intestines, trypsin,

chymotrypsin, elastase & carboxy-

peptidases A & B produce oligopep-

tides in the presence of HCO3- (which

neutralizes the stomach acid &

raises the pH for activation of the

zymogens).

c. Final digestion occurs via amino-

peptidases to produce free AAs and

smaller peptides like di- and

tripeptides.

3. Amino acids are then absorbed by the

intestinal epithelial cells, brought to

the liver (via the portal system) for

metabolism or release into the

general circulation.

ACTIVATION OF THE GASTRIC AND PANCREATIC ZYMOGENS

Proenzymes Active Enzymes

Pepsinogen Pepsin

Trypsinogen Trypsin

Chymotrypsinogen Chymotrypsin

Proelastase Elastase

Procarboxypeptidase A & B Carboxypeptidae A & B

H+ (parietal cells)

Enteropeptidase (enterokinase)

trypsin

trypsin

1. The gastric and pancreatic zymogens involved in protein digestion are

secreted in their inactive precursors or zymogen forms initially and then

converted to their active forms.

2. Pepsinogen, secreted by the chief cells of the stomach, is converted to

pepsin via HCl (from gastric parietal cells) and autocatalytically by pepsin.

3. Trypsinogen is converted to the active form trypsin by enteropeptidase

(formerly enterokinase) from intestinal mucosal cells.

4. Thereafter, trypsin cleaves the other pancreatic zymogens, producing their

active forms.

5. Trypsin is therefore the master or common activator of the pancreatic

zymogens.

trypsin

ACTION OF THE DIGESTIVE

PROTEASES

+

NH3

|

H – C – R

|

C = O

|

NH

|

H – C –R

|

C = O

|

NH

|

H – C – R

|

C = O

|

NH

|

H – C – R

|

C = O

|

NH

|

H – C – R

|

C = O

|

NH

|

H – C – R

|

C = O

|

NH

|

H – C – R

|

COO-

N - terminus

Phe

Tyr

Glu

Asp

Arg

Lys

Phe

Tyr

Glu

Leu

Ala

Gly

Ser

carboxy-

peptidase A

(hydrophobic)

carboxy-

peptidase B

(Arg Lys)

amino-

peptidases

pepsin

trypsin

chymotrypsin

elastase

carboxy-

peptidases

C - terminus

A. Endopeptidases – hydrolyze peptide bonds

at various points within the polypeptide chain.

1. Pepsin preferentially cleaves peptide bonds

in which the carboxyl (carbonyl) group is

provided (or contributed) by aromatic AAs

(Phe, Tyr) and acidic AAs (Glu, Asp).

2. Trypsin specifically cleaves peptide bonds

in which the carboxyl group is provided by

positively charged AAs (Arg, Lys).

3. Chymotrypsin favors hydrolysis of peptide

bonds in which the carboxyl group is provided

by aromatic AAs (Phe,Tyr) and hydrophobic

AAs (Glu, Leu).

4. Elastase cleaves elastin (for which it was

named) and peptide bonds whose carboxyl

group is provided by AAs with small side

chains (Ala, Gly, Ser).

ACTION OF THE DIGESTIVE

PROTEASES

+

NH3

|

H – C – R

|

C = O

|

NH

|

H – C –R

|

C = O

|

NH

|

H – C – R

|

C = O

|

NH

|

H – C – R

|

C = O

|

NH

|

H – C – R

|

C = O

|

NH

|

H – C – R

|

C = O

|

NH

|

H – C – R

|

COO-

N - terminus

Phe

Tyr

Glu

Asp

Arg

Lys

Phe

Tyr

Glu

Leu

Ala

Gly

Ser

carboxy-

peptidase A

(hydrophobic)

carboxy-

peptidase B

(Arg Lys)

amino-

peptidases

pepsin

trypsin

chymotrypsin

elastase

carboxy-

peptidases

C - terminus

B. Exopeptidases – hydrolyze peptide bonds

at outer ends of a polypeptide chain.

1. Carboxypeptidase A – hydrolyzes

peptide bonds at the C-terminus of

hydrophobic AAs (Ala, Ile, Leu, Val).

2. Carboxypeptidase B – hydrolyze peptide

bonds at the C-terminus of Arg and Lys.

3. Aminopeptidases, coming from the

brush border of the epithelial mucosa,

complete protein digestion by

hydrolyzing one amino acid at a time

from the N-terminus of peptides to

produce free amino acids, di- and

tripeptides.

Amino

acid Na+

Amino

acid

Na+

Intestinal

lumen

Brush

border

Amino

acid

Na+

K+

K+

ATP

ADP

+Pi

Active

transporter

Facilitated

transporter

Serosal

side

Portal

vein

TRANSEPITHELIAL AMINO ACID TRANSPORT

1. It is an active process, similar to CHO

digestion in the intestinal epithelial cells.

2. In the brush border membrane is a

semi-specific Na+-dependent transport

protein that carries or pulls with it the AA

as it moves along concentration gradient

(since Na+ conc. in the lumen is > than its

concentration in the inside of the cell).

3. Na+ is then pumped into the serosal side

by the Na+-K+-ATPase system which is

counterbalance by K+ moving in the

opposite direction.

4. The accumulated AA inside the cell is then

transported down its concentration

gradient via facilitated transporters into

the serosal side brought to the liver

via the portal vein for metabolism or for

distribution to other tissues.

Amino

acid Na+

Amino

acid

Na+

Intestinal

lumen

Brush

border

Amino

acid

Na+

K+

K+

ATP

ADP

+Pi

Active

transporter

Facilitated

transporter

Serosal

side

Portal

vein

TRANSEPITHELIAL AMINO ACID TRANSPORT

5. There are different Na+-dependent AA

transport proteins in the apical brush

border membrane of the intestinal

epithelial cells, each transport system

transporting a group of closely related

amino acids:

a. Transport system for neutral AAs.

b. Transport system for proline and

hydroxyproline.

c. Transport system for acidic AAs.

d. Transport system for basic AAs.

(Lys, Arg, Ornithine) and cystine.

6. Some amino acids use facilitated

transport carriers.

7. Most amino acids are transported by more

than one transport system.

9. Ingested dietary amino acids by man are used primarily for synthesis of proteins.

BIOSYNTHESIS

OF THE

NUTRITIONALLY

NON-ESSENTIAL

AMINO ACIDS

CLASSIFICATION OF AMINO ACIDS

Glucogenic

(13 AAs)

Glucogenic

and

Ketogenic

(5 AAs)

Ketogenic

(2 AAs)

Alanine

Asparagine

Aspartate

Cysteine

Glutamate

Glutamine

Glycine

Proline

Serine

Tyrosine

Arginine

Histidine

Methionine

Valine

Isoleucine

Phenylalanine

Tyrptophan

Threonine

Leucine

Lysine

Es

se

nti

al

No

n-e

ss

en

tia

l

P V T T I M H A L L

OVERVIEW OF THE BIOSYNTHESIS OF THE

NON-ESSENTIAL AMINO ACIDS GLUCOSE

3-Phosphoglycerate

Pyruvate Alanine

Serine

Glycine

Cysteine

Methionine

Acetyl CoA

Oxaloacetate Aspartate

Asparagine

Glutamine Citrate

Isocitrate

-ketoglutarate Glutamate

Glutamine

Proline

Arginine

Phenylalanine Tyrosine

1. There are 11 non-essential amino acids of the 21common amino acids.

2. Non-essential because they can be synthesized in sufficient amounts by the body from:

a. Amphibolic intermediates of:

a1. Glycolysis

i. 3-phosphoglycerate - Cysteine, Serine, and Glycine

ii. Pyruvate - Alanine

a2. Citric Acid Cycle

i. -ketoglutarate – Glutamine, Glutamate, Proline and Arginine

ii. Oxaloacetate – Aspartate and Asparagine

b. Phenylalanine – Tyrosine

Glutamate

I. AMINO ACIDS DERIVED FROM GLYCOLYSIS

3-phosphoglycerate dehydrogenase

PLP

NAD+ NADH

+ H+

α-Ketoglutarate

Pi

Phosphoserine phosphatase

Glycolysis

Serine

A. Serine C1OO-

I

H – C2 – OH

I

C3H2

I

O - PO32-

3-Phosphoglycerate

C1OO-

I

C2 = O

I

C3H2

I

O - PO32-

3-Phosphohydroxypyruvate

COO-

|

H3N+ - C – H

|

CH2

|

CH2

|

COO-

Glutamate

C1OO-

I

H3N – C2 – H

I

C3H2

I

O - PO32-

3-Phosphoserine

+

C1OO-

I

H3N – C2 – H

I

C3H2

I

OH

+

Glucose

aminotransferase

(keto acid)

Glycine D- serine

(CNS neurotransmitters) Selenocysteine

1. 3-phosphoglycerate (from

glycolysis) is converted to 3-phos-

phohydroxypyruvate, a keto acid, via

3-phosphoglycerate dehydrogenase

with NAD+ as a hydrogen acceptor

reduced to NADH2.

2. Followed by transamination with

glutamate as the amino group donor

to 3-phosphohydroxypyruvate,

forming 3-phosphoserine and -

ketoglutarate in the presence of

PLP,a cofactor, via an

aminotransferase.

3. Finally, phosphoserine phosphatase

hydrolyzes the phosphate group

(dephosphorylation) to form serine.

4. Serine is a precursor of the neuro-

transmitters glycine and D-serine;

also a component of the unusual AA

selenocysteine found in glutathione

peroxidase.


Other Pathway For Serine

H2C – NH3+

|

COO-

Glycine

CH2OH

|

H – C – NH3+

|

COO-

Serine N5, N10 – CH2 – TH4 (N5-N10-Methylene THF)

TH4

(Tetrahydrofolate)

Serine hydroxymethyl

transferase

PLP

H20

H20

1. Serine may also be formed reversibly from glycine by transfer of a

hydroxymethyl group via serine hydroxymethyl transferase in the

presence of PLP and tetrahydrofolate (TH4), forming N5, N10-Methylene

TH4.

2. The demand for serine and glycine and the amount of N5, N10-Methylene

TH4 determine the direction of the reaction.

I. AMINO ACIDS DERIVED ROM GLYCOLYSIS

COO-

I

H3N – C – H

I

CH2OH

Serine

COO-

I

H3N – C – H

I

H

Glycine

+ +

5

H

C

C N

CH2

I

N –

H

H

CH2

N H

10 Tetrahydrofolate

(FH4)

N5-N10-Methylenetetrahydrofolate

(N5-N10-CH2-H4 folate)

Serine hydroxymethyl transferase

PLP

Methylene

5

H

C

C N

CH2

I

N –

H

CH2

N

10 I

H2C

Reactive part of tetrahydrofolate

7

6

9

8

3-Phospho-

glycerate

1. Serine, coming from the glycolytic intermediate 3-phosphoglyerate, is converted reversibly to

glycine via serine hydroxymethyltransferase, with PLP as a coenzyme and tetrahydrofolate (FH4).

2. The reaction involves the transfer of a one-carbon unit from serine with tetrahydrofolate as an

acceptor.

3. FH4 is converted to N5-N10-methylenetetrahydrofolate (methylene, a one-carbon unit is bound to 2 of

the nitrogens of the carrier molecule).

4. Hence FH4 is a carrier of one-carbon units (-CH2) in metabolic pathways; the other carriers are

biotin, a carrier of CO2 in carboxylation reactions and S-adenosylmethionine (SAM), a carrier of

methyl groups (-CH3) in methylation reactions.

B. Glycine – the Major Pathway


B. Glycine

H2C – NH3+

|

COO-

Glycine

PLP

CH3 H

| |

H – C – C – C

| |

OH NH4+

Threonine

O-

O

Minor Pathway

Threonine is degraded into glycine in

the presence of PLP s a cofactor.


C. Cysteine CH2 – OH

|

H – C – NH3+

|

COO-

Serine

SH

|

CH2

|

CH2

|

H – C – NH3+

|

COO-

Homocysteine

-OOC – CH – CH2 – S

| |

+NH3 CH2

|

CH2

| +

H – C – NH3

|

COO-

Cystathionine

-OOC – CH – CH2 – SH

+|

NH3

H2O Cystathione -synthase PLP

H2O

NH4+

PLP

α-Ketobutyrate

Methionine

-Cystathionase

Cysteine

COO-

I

H3N – C – H

I

CH2

I

CH2

I

H3C - S

+

Serine Methio-

nine

Cystathionuria

def

ATP

S-adenosyl

methionine

(SAM; Adomet)

S-adenosyl

homocysteine

(SAH)

CH3

adenosine

Methionine adenosyltransferase

Adenosylhomo- cysteinase

Methylase

1. Methionine (an essential AA) condenses with ATP via

methionine adenosyltransferase to form S-adenosylmethio-

nine (SAM; Adomet); methyl group of SAM is cleaved by

methyltransferase to form S-adenosylhomocysteine (SAH);

Adenosine group of SAH is cleaved by adenosylhomocys

teinase to form homocysteine.

2. Serine condenses with homocysteine (from methionine) to

form cystathionine via cystathione -synthase with PLP as a

co-factor.



|

H – C – NH3+

|

COO-

Serine

SH

|

CH2

|

CH2

|

H – C – NH3+

|

COO-

Homocysteine


| |

+NH3 CH2

|

CH2

| +

H – C – NH3

|

COO-

Cystathionine


+|

NH3


H2O

NH4+

PLP

α-Ketobutyrate

Methionine

-Cystathionase

Cysteine

COO-

I

H3N – C – H

I

CH2

I

CH2

I

H3C - S

+

Serine Methio-

nine

Cystathionuria

def

ATP

S-adenosyl

methionine

(SAM; Adomet)

S-adenosyl

homocysteine

(SAH)

CH3

adenosine



Methylase

3. Cystathione is then cleaved by PLP-dependent -cystathio-

nase into -ketobutyrate (and NH4+) and cysteine.

4. Serine contributes the carbons and nitrogens for cysteine;

methionine provides the sulfur of cysteine via transulfuration

- one of the routes used for methionine catabolism.

5. Because methionine is an essential amino acid, cysteine

synthesis can be sustained only if the dietary intake of

methionine is adequate.



|

H – C – NH3+

|

COO-

Serine

SH

|

CH2

|

CH2

|

H – C – NH3+

|

COO-

Homocysteine


| |

+NH3 CH2

|

CH2

| +

H – C – NH3

|

COO-

Cystathionine


+|

NH3


H2O

NH4+

PLP

α-Ketobutyrate

Methionine

-Cystathionase

Cysteine

COO-

I

H3N – C – H

I

CH2

I

CH2

I

H3C - S

+

Serine Methio-

nine

Cystathionuria

def

ATP

S-adenosyl

methionine

(SAM; Adomet)

S-adenosyl

homocysteine

(SAH)

CH3

adenosine



Methylase

6. A genetic deficiency of cystathione -synthase or

PLP causes cystathionuria or the presence of

cystathione in the urine; a benign disorder with no

clinical abnormalities.

II. AMINO ACIDS RELATED TO TCA INTERMEDIATES

1. Glutamate

A. Amino Acids Related to α-ketoglutarate

COO-

|

C = O

|

CH2

|

CH2

|

COO-

α-ketoglutarate

COO-

|

H3N+ - C – H

|

CH2

|

CH2

|

COO-

Glutamate

NAD(P)+ NAD(P)H

NH4+

Glutamate dehydrogenase

1. Reductive amination of -ketoglutarate to glutamate via glutamate dehydrogenase

is a freely reversible reaction; NAD+ or NADP is a cofactor.

2. The 5-carbons of glutamate came from -ketoglutarate which also came from glucose.

3. Glutamate is used for the synthesis of other amino acids: glutamine, proline,

arginine, ornithine and glutathione (an antioxidant).

Synthesis of other AAs

Glutamine, Proline

Arginine, Ornithine

Glutathione

SH

I

CH2

I

HC – NH3

I

COO-

+

Cysteine

ATP ADP + Pi

-Glutamylcysteine COO-

I

H3N – C – H

I

H

ATP

ADP + Pi Glycine

Glutathione (-Glutamylcysteinylglycine)

GLUTAMATE AS A

PRECURSOR OF

GLUTATHIONE

-carbon or

3rd carbon

Sulfhydryl group of cysteine

SH

I

CH2

I

CH – C – N – CH2

I

COO-

H

O

II

C – N –

I

C3H2

I

C2H2

I

HC1 – NH3

I

COO-

H

+

O

II

COO-

|

H3N+ - C – H

|

CH2

|

CH2

|

COO-

Glutamate

- carboxyl group

- carboxyl group

-glutamylcysteine synthase

Glutathione synthase

1. Glutamate, in the presence of

ATP, condenses with cysteine

to form the dipeptide -gluta-

mylcysteine.

2. Followed by the addition of

glycine, again in the presence

of ATP to form the tripeptide -

glutamylcysteinylglycine; the

letter refers to the 3rd carbon

in the molecule, counting the

one bonded to the amino group

as the 1st carbon (recall that

is the 3rd letter in the Greek

alphabet).

GLUTATHIONE AS AN ANTIOXIDANT

Sulfhydryl

group

2 - Glu – Cys - Gly - Glu – Cys -Gly

I

SH

Glutathione

(GSH; reduced form)

I

S

I

S

I

- Glu – Cys - Gly

Glutathione disulfide

(GSSG; oxidized form)

2 H2O

H2O2

Glutathione peroxidase

2 GSH

(reduced form)

NADPH

+ H+

NADP+

Glutathione reductase

Disulfide bond

2 - Glu – Cys - Gly I

SH

1. The sulfhydryl group of glutathione

(GSH; reduced form) reduces or

scavenges H2O2 (an oxidizing agent)

to H2O, catalyzed by glutathione

peroxidase, forming glutathione

disulfide (GSSH; oxidized form)

consisting of 2 molecules of the

reduced form joined together by a

disulfide bond between the –SH

groups of the 2 cysteine residues.

2. Reduced glutathione (GSH) is

regenerated via glutathione

reductase at the expense of NADPH.

GLUTATHIONE IS INVOLVED IN THE TRANSPORT OF AMINO

ACIDS ACROSS PLASMA MEMBRANES Amino acid

-Glutamyl transpeptidase

-Glu-Cys-Gly Cys-Gly -Glu-Amino acid Outside

Inside Cys-Gly

Cysteine +

Glycine

H2O -Glutamyl

cyclotransferase

ATP + H2O 5-oxopro-

linase

5-Oxoproline

L-Glutamate -Glutamylcysteine

synthase

ADP + Pi

+ 2 H+

ADP + Pi

+ 2 H+

Plasma membrane

-Glu-Amino acid

-Glutamyl- cysteine Glutathione

synthase

Amino

acid

ATP

-Glu-Cys-Gly

(glutathione)

-Glu-Cys-Gly

(glutathione)

1. Transport of amino acids uses the -glutamyl cycle (Meister Cycle).

2. Membrane-bound -glutamyltranspeptidase catalyzes the transpeptidation of -

glutamyl residue from glutathione to the amino acid, forming -glutamyl-amino acid

and cysteinylglycine, the remaining portion of glutathione.

3. The -glutamyl-amino acid (a dipeptide, consisting of 2 amino acids glutamate and the

amino acid to be transported) is transported into the cytoplasm to form 5-oxoproline.

4. Once inside the cell, the cytoplasmic enzyme -glutamylcyclotransferase releases the

amino acid and cyclizes the -glutamyl group to form 5-oxoproline, which is converted

to glutamate via 5-oxoprolinase in the presence of ATP.

GLUTATHIONE IS INVOLVED IN THE TRANSPORT OF AMINO

ACIDS ACROSS PLASMA MEMBRANES

Amino acid

-Glutamyl transpeptidase

-Glu-Cys-Gly Cys-Gly -Glu-Amino acid Outside

Inside Cys-Gly

Cysteine +

Glycine

H2O -Glutamyl

cyclotransferase

ATP + H2O 5-oxopro-

linase

5-Oxoproline

L-Glutamate -Glutamylcysteine

synthase

ADP + Pi

+ 2 H+

ADP + Pi

+ 2 H+

Plasma membrane

-Glu-Amino acid

-Glutamyl- cysteine Glutathione

synthase

Amino

acid

ATP

-Glu-Cys-Gly

(glutathione)

-Glu-Cys-Gly

(glutathione)

5. To complete the -glutamyl cycle, cysteine condenses with glutamate via

-glutamylcysteine synthase, again in the presence of ATP, forming

-glutamylcysteine.

6. -glutamylcysteine then condenses with glycine via glutathione synthase

again in the presence of ATP, forming and resynthesizing glutathione.


2. Glutamine


NH2

|

C = O

|

CH2

|

H – C – NH3+

|

COO-

Glutamine

COO-

|

H3N+ - C – H

|

CH2

|

CH2

|

COO-

Glutamate

NH4+

ATP ADP + Pi

Glutamine synthase

H2O NH4

+

Glutaminase

1. Glutamine coming from transamination of α-ketoglutarate, is aminated to glutamnine

via glutamine synthase by adding free NH4+ (as the amino group donor) to the

carboxyl group (-COOH) of the side chain of glutamate in the presence of ATP.

2. In the presence of glutaminase, glutamine is deaminated to glutamate.

3. Asparagine is the other amino acid that is synthesized from amination of aspartate.

Α-ketoglutarate


3. Proline


H - C – CH2 – CH2 – CH – COO-

II

O

Proline

-COO – CH2 – CH2 – CH – COO-

Glutamate

Glutamate semialdehyde

CH2 H2C

CH – COO- HC +

N

Spontaneous cyclization

NADH +

H+

NAD+

NADH

+ H+

NAD+

ATP

ADP + Pi

NH3

+

NH3

+

Δ1-Pyrroline 5-caboxylate H

+

N

CH2 H2C

CH – COO- H2C

H2

FAD . 2H

FAD

NADPH

+

H+

NADP+

Pyrroline-5-

carboxylate

reductase

1. Glutamate is phosphory-

lated by ATP and then

converted to glutamate

semialdehyde via the

reduction of side chain

carboxyl group (-COOH)

to an aldehyde.

2. Followed by

spontaneous

cyclization of to Δ1-

pyrroline 5-carboxylate.

3. This cyclic compound is

then reduced by NADPH

to proline via pyroline-5-

carboxylate reductase.


4. Arginine


Glutamate

Arginine

+

NH3

|

H – C – CH2 – CH2 – CH – COO-

||

O

Glutamate semialdehyde

+

NH3

|

H3N+ - CH2 – CH2 – CH2 – CH – COO-

Transanimation Ornithine aminotransferase

Ornithine

+

NH NH3

|| |

H2N – C – CH2 – CH2 – CH2 – CH – COO-

Urea

cycle

-Ketoglutarate 1. Glutamate, coming from

-ketoglutarate, is

converted to glutamate

semialdehyde.

2. Followed by transamina-

tion into ornithine (an

intermediate of the urea

cycle) arginine.

3. The amounts of arginine

generated by the urea

cycle is adequate only

for the adult and are

insufficient for growth,

hence during periods of

rapid growth as in

infancy and childhood,

arginine becomes an

essential amino acid.


B. Amino Acids Related to Oxaloacetate

O

||

C – NH2

|

CH2

|

H – C – NH3+

|

COO-

Asparagine

Aspartate & Asparagine

COO-

|

CH2

|

H – C – NH3+

|

COO-

Aspartate Glutamine ATP

AMP + PPi

Glutamate

Asparagine synthetase

NH4+

H2O

Asparaginase

COO-

|

CH2

|

C = O

|

COO-

Oxaloacetate

PLP

Transanimation

1. Oxaloacetate is transami-

nated to aspartate with PLP

as a cofactor in a freely

reversible reaction.

2. Aspartate is then aminated to

asparagine via asparagine

synthetase in the presence of

ATP and glutamine as a

source of nitrogen (or the

amino group donor).

3. Leukemic cells require

asparagine for their growth,

hence asparaginase has been

used as an anti-tumor agent.

It acts by converting

asparagine to aspartate in the

blood, thereby decreasing the

amount of asparagine

available for tumor growth.

III. TYROSINE COMING FROM PHENYLALANINE

Quinonoid dihydrobiopterin (BH2)

- CH2–CH–COO-

NAD+

NADH

+ H+

H2N N H

N H

H

HN

II

O

N

H

H

CH – CH – CH3

I

OH

I

OH

H2N N H

N H

H

N

II

O

N

H

CH – CH – CH3

I

OH

I

OH

+

NH3

I HO -

O2

H2O

- CH2–CH–COO-

+

NH3

I

Phenylalanine

Tyrosine

Tetrahydrobiopterin

(BH4)

Phenylalanine hydroxylase

GTP Biosynthesis

Dihydropteridine reductase

2 1

5 4 3 6

8

9

7

10

3 5

1. Phenylalanine is reduced to tyrosine via phenylalanine hydroxylase (PAH), a mixed-

enzyme oxidase with 2 activities

a. Activity I: Reduction of O2 to H2O and of phenylalanine to tyrosine (molecular O2

donates one atom to H2O and one atom to the product tyrosine).



- CH2–CH–COO-

NAD+

NADH

+ H+

H2N N H

N H

H

HN

II

O

N

H

H

CH – CH – CH3

I

OH

I

OH

H2N N H

N H

H

N

II

O

N

H

CH – CH – CH3

I

OH

I

OH

+

NH3

I HO -

O2

H2O

- CH2–CH–COO-

+

NH3

I

Phenylalanine

Tyrosine

Tetrahydrobiopterin

(BH4)


GTP Biosynthesis


2 1

5 4 3 6

8

9

7

10

3 5

b. Activity II: Reduction of the co-factor tetrahydrobiopterin (BH4) to dihydrobiopterin (BH2); the H

atoms on carbons 3 and 5 of BH4 are absent in BH2.

2. BH4 is synthesized in the body from GTP; BH4 is usually used in the hydroxylation of aromatic amino

acids like phenylalanine, tyrosine and tryptophan (with aromatic rings).

3. BH2 is reduced back to BH4 via an NADH-dependent dihydrobiopterin reductase for phenylalanine to

continue forming tyrosine.



- CH2–CH–COO-

NAD+

NADH

+ H+

H2N N H

N H

H

HN

II

O

N

H

H

CH – CH – CH3

I

OH

I

OH

H2N N H

N H

H

N

II

O

N

H

CH – CH – CH3

I

OH

I

OH

+

NH3

I HO -

O2

H2O

- CH2–CH–COO-

+

NH3

I

Phenylalanine

Tyrosine

Tetrahydrobiopterin

(BH4)


GTP Biosynthesis


2 1

5 4 3 6

8

9

7

10

3 5

4. Tyrosine is unique because unlike all the other non-essential AAs which are

synthesized from certain intermediates of glycolysis (3-phospholycerate and pyruvate)

and Krebs Cycle, it results from a simple one-step hydroxylation of phenylalanine.

5. Hence the presence of dietary tyrosine decreases the need for phenylalanine.

IV. BIOSYNTHESIS OF HYDROXYPROLINE

AND HYDROXYLYSINE

18O2

Ascorbate

Fe2+

Prolyl hydroxylase (Lysyl hydroxylase)

Pro (Lys)

18OH

Pro (Lys)

α-ketoglutarate [18O] Succinate

COO-

I

CH2

I

CH2

I

C = O

I

COO-

COO-

I

CH2

I

CH2

I

COO-

1. Peptide-bound proline and lysine are hydroxylated by prolyl hydroxylase and lysyl

hydroxylase respectively (found in skin and skeletal muscles, including granulating

wounds), in the presence of molecular O2 (one atom of O2 is incorporated into

proline or lysine; the other atom into succinate), ascorbate , Fe+2 and -ketoglutarate.

2. Hydroxyproline and hydroxylysine are present principally in collagen; collagen is a

glycoprotein in all tissues and organs provides the framework that gives tissues

their form and structure.

3. A deficiency of Vitamin C or hydroxylase results to scurvy.

Amino acids and protein chemistry 1

Health & Medicine

Transcript of Amino acids and protein chemistry 1