AMINO ACIDS IONIZATION STATES AND ISOELECTRIC POINT

Question 1

___ can gain or lose protons (H+) depending on the pH of the environment, which affects their charge

Answer 1

Amino acids

Question 2

Each amino acid had at least two ionizable groups:

• ___ (-NH3+) BASIC, can lose proton
• ___ (-COO): ACIDIC, can gain a proton

Answer 2

Amino group
Carboxyl group

Question 3

IONIZATION STATES AT DIFFERENT pH levels

• ____ (acidic) : All groups are protonated –> net ____ charge
• Neutral pH: Zwitterion form → net charge = ___
• High pH (basic): All groups are deprotonated –> net ___ charge

Answer 3

low pH
Positive
0
negative

Question 4

the pH at which the amino acid has no net charge (i.e., it’s a zwitterion).

Answer 4

Isoelectric point

Question 5

How to calculate the isoelectric point (pl)

Answer 5

Pl= pKa COOH + pKa NH3+ /2

Question 6

Protein purification: Techniques like isoelectric focusing rely on pI.

Solubility: ____ are least soluble at their pI.

Charge interactions: pI affects how proteins and amino acids
interact with each other and with membranes.

Answer 6

Amino acids
Pl

Question 7

Example of pl (amino acids)

1. Glycine
   • pKa (COOH) = 2.3
   • pKa (NH3+) = 9.6
   • pl = (2.3 +9.6)/2 = 5.95
   • Zwitterion at pH ≈ 6 2
2. Alanine
   PI = 2.3 + 9.7 /2 = 12.0/ 2
   pl= 6.0

Question 8

Examples :
• These amino acids have three ionizable groups, so you must consider the side chain pKa too
1. Glutamic acid (Glu)
• pKa (COOH) = 2.2
• pKa (R group, -COOH) = 4.3
• pKa (NH3+) = 9.7
• pl= (2.2. + 4.3) /2 = 3.25

Question 9

When calculating the isoelectronic point (pl) of an amino acid with three ionizable groups, we only use the two pKa values that surround the zwitterionic form– the form where the molecule HAS NO NET CHARGE

pl= 2.2 + 4.3 /2 = 6.5 /2 = 3.25

Question 10

Zwitterion exits between the two acidic forms

• at pH - 3.25, the main COOH is depronated (-COOH), the R-group COOH is still protonated, and NH3+ is still positive —> net charge = 0

pl= 2.2 + 4.3 /2 = 3.25

• We do not use the NH3+ pka because by the time it deprotonates (at pH -9.7) the molecule is already negatively charged

Question 11

EXAMPLE

• you average the two basic pKa values because Zwitterion exists between the two positive states

2. Lysine (lys)
   • pKa (COOH) = 2.2
   • pKa (NH3+) = 9.0
   • pKa (R group, -NH3+) = 10.5
   • Pl = (9.0 + 10.5)/2 = 9.75

Question 12

___ = a hydrogen ion (H⁺)

___ = losing a proton (H⁺)

Answer 12

Proton
Depronated

Question 13

• a ring-shaped group found in the side chain of the amino acid histidine.

• It can gain or lose a proton, which makes it important in enzyme reactions.

Answer 13

Imidazole is a ring

Question 14

Think of imidazole like a switch on a machine:

• When it has a proton, it’s on (positively charged).

• When it loses the proton, it’s off (neutral).

This switch helps enzymes start or stop chemical reactions.

Question 15

___ a molecule that has:

• Both a positive and a negative charge

• But the overall charge is zero

• Imagine a magnet with a + on one end and a – on the other.
It has charges, but it doesn’t move because the forces cancel out.

Answer 15

Zwitterion

Question 16

Amino acids in your body mostly exist as zwitterions at normal pH ___

• They help proteins fold correctly and interact with water.

• Zwitterions are STABLE, SOLUBLE, AND REACTIVE.

Answer 16

-7.4

Question 17

IONIZATION STATE

Alanine :
• ___ side chain type
• ___ pl (approx)
• Simple, no IONIZABLE R group

Answer 17

Non polar
-6.0

Question 18

IONIZATION STATE

Histidine:
• ___ side chain type
• Pl approx: ___
• IMIDAZOLE side chain (pKa -6.0)

Answer 18

Polar, basic
-7.6

Question 19

IONIZATION STATE

• side chain type : BASIC
• pl= ____
• Guanidino group (pKa -12.5)

Answer 19

Arginine
-10.75

Question 20

IONIZATION STATE

Side chain type: ACIDIC
pl approx : ___

• EXTRA CARBOXYL GROUP (pKa -3.9)

Answer 20

Aspartic acid
-2.9

Question 21

IONIZATION STATE

Tyrosine
Side chain type: ___,___
Pl approx:

• PHENOL GROUP (pKa -10.1)

Answer 21

Polar
Aromatic
-5.7

Question 22

___: pH from 0 to 14.

___: Net charge of the amino acid.

• Where the curve crosses __: That’s the isoelectric point (pI)—the pH at which the amino acid has no net charge.

• Steep drops or rises: Indicate ionization of specific
groups (carboxyl, amino, or side chain).

Answer 22

X-axis
Y-axis
0

Question 23

ELECTROPHORESIS & PROTEIN SEPARATION

1. If pH < pI–> Amino acid is ___ —> Moves to the negative electrode.
2. If pH > pI–> Amino acid is ___ —> Moves to the positive electrode.

At pI –> No movement (used in ISOELECTRIC FOCUSING).

Answer 23

+ charged
– charged

Question 24

• DRUG SOLUBILITY
If a drug contains ___, its charge affects HOW WELL IT DISSOLVES in blood (pH ~7.4).

• ENZYME ACTIVITY
Enzymes work best at certain pH levels because their __ depend on ionization states.

Answer 24

Amino acids
active sites

Question 25

Imagine an amino acid as a person holding two balloons: One balloon is acidic (–COOH)--> Can let go of a H⁺ (like releasing a balloon). The other balloon is basic (–NH₂) ---> Can grab a H⁺ (like catching a new balloon).

Question 26

Ionization State - Analogy Depending on the pH (how acidic or basic the environment is), this person will either: Hold both balloons (____). Release one balloon (___) Release both balloons (___)

Answer 26

fully protonated, + charge zwitterion, neutral). fully deprotonated, – charge).