BCMB 3100 - Lecture 3 Horton Chapter 3

Know the structure, names and classes of the 20 common amino acids Know pKas of -amino and carboxyl groups, and any ionizable side groups Know potential H-bonding and salt-bridging interactions Titration curve of amino acids
(Problems: 4,6,8,9,10,15)

Overview
Diversity of protein function Definition of amino acids Memorize complete structure of 20 common amino acids!!! pKas of amino and carboxyl groups Amino acids with ionizable side groups Titration curve

Examples of Protein Function

Enzymatic catalysis: enzymes increase reaction rates by 106-fold; nearly all known enzymes are proteins dictates chemistry and metabolism Transport & storage: small molecules and ions are transported by proteins - examples: hemoglobin, myoglobin Coordinated motion: examples: myosin and actin (muscle movement)

Examples of Protein Function

Mechanical support: Immune protection:
antibodies collagen (tensile strength to skin and bones)

Generation & transmission of nerve impulses: Control of growth and differentiation:

acetylcholine receptor DNA binding proteins (repressors, activators, transcription factors, hormones, regulation of translation)

Protein Function - Catalysis

Catalysts: enzymes increase reaction rates
Urease

urea

G ~ 25 kcal mol-1 or 104.5 kJ mol-1

Test tube - 2,642 yrs/molecule Vs Cell - 37 sec/molecule

Rate enhancement of 1015

James Sumner
Nobel Prize 1946

Showed enzymes are proteins

Protein Function - Transport

Many small molecules and ions are transported by proteins.
Examples: hemoglobin, transferrin, pores.

Transferrin

Fig. 4.49, Horton

Hemoglobin
Fig .49

Protein Function Coordinated Motion

Motion: Combination of enzymes with structural proteins. Examples: flagella, or muscle (myosin and actin).

Fig1.14

Protein Function Mechanical Support

Structural proteins shapes the cells (tubulin and actin) and gives tensile strength to skin (collagen).

Tubulin - GFP collagen triple helix

actin

Protein Function Gene Expression

Control of cell information such as gene expression and mRNA translation (eg. repressors and activators).

Fig 21.14

Protein Function Cell Signaling

Cytokines, hormones, receptors Examples: Insulin, leptin, HGH.

Signal
Fig 9.46

Protein Function Specialized functions

Examples: toxins and antibodies.

Anthrax toxin PA
Fig 4.54

WHAT ARE PROTEINS?

Proteins are macromolecules made up of amino acids (20 amino acids) -amino acids: amino group, carboxyl group, a hydrogen atom and a distinctive R group R group is bonded to the -carbon because it is adjacent to the carboxyl group.

Amino acid nomenclature

O C
+H 3N

O-

CH2

OH
Ball and stick 3D representation

Side chain position denoted by Greek letters - , , , , , , (alternate = a, b, g, d, e, z, h

Amino acids are charged

Amino acids in solution will be in a charged state -amino group and/or the -carboxyl group will be charged depending upon the pH The R group may also be charged

Under normal cellular conditions amino acids are zwitterions (dipolar ions): Amino group = Carboxyl group = -NH3+ -COO-

General Structure of the ionized from of an amino acid

COO-

NH3+ C H

R
COO-: pKa = 1.8 - 2.5
NH3+: pKa = 8.7 - 10.7
Fischer Projection

You MUST know this!!!

Amino acids are chiral

Amino acids are chiral (asymmetric) 4 different groups attached to the -carbon Glycine is the exception (R = H) Enantiomers of amino acids are called D (right-handed) or L (left-handed) L and D refer to absolute configuration L-amino acids are the only constituents of proteins
Fig 3.2

Amino acid side chains

20 different R groups (side chains) that differ in
Size Shape Charge hydrogen-bonding capacity chemical reactivity

Sequence of amino acids in protein gives the protein its All biochemists must remember
Shape Function

Structure of each amino acid Three letter code for each amino acid Single letter code for each amino acid

Structures of the 20 Common Amino Acids

Classes of amino acids: Aliphatic Aromatic Sulfur-containing Alcohols Bases Acids Amides You must be able to draw the structure of the 20 amino acids in their correct ionized form at any given pH
B A S I C
O
+

O
+

O
+

O
+H 3N

H3N CH CH3

C O-

H3N CH CH H3C

C O-

H3N CH

C O-

C CH CH O-

Alanine Ala A

H3C
CH3

CH2 CH CH3

Aliphatic
O
+H N 3

Valine Val V
O

Leucine Leu L
O

H2C CH3

CH3

Isoleucine Ile I
O

C CH O-

H2 + N

C O-

+H

3N

C CH CH2 O-

+H N 3

C CH CH2 O-

Glycine Gly G

Aliphatic/Structural
O
+

Proline Pro P

HS

Cysteine Cys C
H3C

H2C S

O
+H 3N

Sulfurcontaining
O
+

Methionine Met M
O C CH CH2 O-

H3N CH

C O
-

C CH CH2 O-

H3N CH

C O-

H3N

CH2

HN

Histidine His H
O

H2C C O O-

Glutamate Glu E

O C O

CH2

Acidic

Aspartate Asp D

Phenylalanine Phe F
O
+H N 3

O
+

O
+H 3N

H3N CH CH2 H2C CH2

C O+H N 3

C CH CH2 O-

C CH O-

C CH CH2 O-

H2C NH3+

Lysine Lys K
O

H2C C O NH2

Glutamine Gln Q

H2N C O

CH2

Asparagine Asn N

HO

Tyrosine Tyr Y
O
+H 3N

Amide
OO
+

+H

3N

C CH CH2

A R O M A T I C

C CH CH2 O-

O
+

H2C CH2
+HN

H3N CH CH2 HO

C O-

H3N CH CH H3C

C O-

HN
OH

C H2N NH2

Arginine Arg R

Serine Ser S

Threonine Thr T

Alcohol

Tryptophan Trp W

The aliphatic amino acids

Aliphatic Side chains contain only Cs and Hs Glycine not chiral (R = H)

P. 57

Isoleucine (Ile)
Ile has 2 chiral carbons, 4 possible stereoisomers

P. 57

Proline (Pro)
Proline (Pro, P) - has a three carbon side chain bonded to the a-amino nitrogen The heterocyclic pyrrolidine ring restricts the geometry of polypeptides

P. 58

The Aromatic amino acids

pKas = 10.5

benzene ring (260 nm)

phenol ring (278 nm)

indole group (280 nm) P. 58

(UV absorbance)

UV Absorbance

Tyr

BSA (bovine serum albumin is a protein that has Trp, Tyr and Phe amino acids

Sulfur containing amino acids

pKas = 8.4

P. 59

Formation of cystine (disulfide bond)

(2 e- Oxidation)

O
+H 3N

H- (H with an extra e-) NOT H+

O-

O
+H 3N

C CH CH2

Oxidation

R-X
R-XH2

C CH CH2 O-

HS

+

H2C
-O

S
SH

+ 2H+ + 2e-

R-X

R-XH2

-O

H2C CH C O NH3+

CH C O NH3+

Reduction

Cysteine

Reducing conditions in the cell are due to Glutathione (GSH). GSH is a tri-peptide (ECG) with a reducing thiol. See problem 6 in the book.

Cystine or Disulde

Cleaving a disulfide bond)

O
+H 3N

C CH CH2 O-

-MercaptoEthanol
(BME)

+H

3N

C CH CH2 O-

S S H2C
-O

H2C
-S

OH

Disulde

H2C
-O

CH2

CH C O NH3+

CH C O NH3+

BME

OH
+H 3N

C CH CH2 O-

OH

Cys

C H2

H2 C SS S

H2C CH2
+H 3N

O C CH CH2
-S

-S

O-

HO CH2 H2C S S H2C CH2

OH
-O

H2C CH C O NH3+

Amino acids containing alcohol groups

Serine (Ser, S) and Threonine (Thr, T) have uncharged polar side chains

*
*

* Chiral center

2 chiral centers

P. 59

Basic Amino acids

pKas = 6.0

pKas = 10.5

pKas = 12.5

imidazole

alkylamino group

guanidino group

P. 60

Acidic amino acids and their amides

pKas = 3.9

pKa = 4.1

dicarboxylic acids

uncharged but highly polar

P. 60

Hydrophobicity of Amino Acid Side Chains

Hydropathy: the relative hydrophobicity of each amino acid The larger the hydropathy, the greater the tendency of an amino acid to prefer a hydrophobic environment Hydropathy affects protein folding:

hydrophobic side chains tend to be in the interior hydrophilic residues tend to be on the surface

Table 3.1 Hydropathy scale for amino acid residues

Hydropathy: the relative hydrophobicity of each amino acid apolar solvent
G = -RT ln Keq

Amino acid

Keq =

[Aqueous] [Apolar]

water

Highly hydrophobic
Isoleucine

Phenylalanine
Valine

Leucine

Methionine
Less hydrophobic
Tryptophan
Alanine

Glycine

Cysteine

Tyrosine

Proline

Threonine

Serine

Highly hydrophilic
Histidine

Glutamate
Asparagine
Glutamine
Apartate

Lysine

Arginine

Free-energy change for transfer (kjmol-1)

3.1

2.5

2.3

2.2

1.1

1.5*

1.0

0.67

0.17

0.08

-0.29

-0.75

-1.1

-1.7

-2.6

-2.7

-2.9

-3.0

-4.6

-7.5

Amino acids are charged

-amino group and/or the -carboxyl group will be charged depending upon the pH. The R group may also be charged. At neutral pH amino acids are predominantly dipolar ions (zwitterions). (NH3+ NH2 + H+ ) NH3+ ( COOH COO- + H+ ) COO-

C H

Note: pH dependence R

of the ionization state

COO-: pKa 1.8 - 2.5 NH+: pKa 8.7-10.7

Ionization of Amino Acids

Ionizable groups in amino acids: (1) -carboxyl, (2) -amino, (3) some side chains Each ionizable group has a specific pKa AH B + H+ For a solution pH below the pKa, the protonated form predominates (AH) For a solution pH above the pKa, the unprotonated (conjugate base) form predominates (B)

Titrating a zwitter ion (2 pKas)

+1
OH
C C CH3 H

pKa
(COOH)

H+

+H 3N

0
OC C CH3 H

pKa
(NH3)

H+

H2N

-1
OC C CH3 H

+H

3N

H+

H+

Increasing pH

Titration curve for Alanine

Titration curves are used to determine pKa values pK1 = 2.4 pK2 = 9.9 pIAla =
isoelectric point

pI = pH where the net charge is 0
pI = (2.4+9.9) / 2 = 6.15

P. 63

pKa values of amino acids

The R group of 7 amino acids are also ionizable The pKa of these R groups can range from 3.9 to 12.5 You must memorize the pKas of the side chains of these seven amino acids!
The ratio of unprotonated to protonated R-group changes by an order of magnitude with each change in pH unit relative to the groups pKa. P. 64

Ionization of Histidine
(a) Titration curve of histidine pK1 = 1.8 pK2 = 6.0 pK3 = 9.3 pI = pH at which net charge is zero

Lehninger Principles of Biochemistry, 3rd ed.

To determine the pI: pick the pKa above & below point where charge is zero and average those 2 pKas

P. 64

Deprotonation of imidazolium ring

pKa1 = 1.8

pKa3 = 9.3

Net charge +1

Net charge 0
Fig 3.7

pI = (6.0 +9.3)/2 = 7.65

The dipeptide sweetner aspartame

Asp

Phe

CH3

Aspartame is a dipeptide methyl ester (aspartylphenylalanine methyl ester) About 200 times sweeter than table sugar Used in diet drinks
Fig 3.10