Biochemistry: Protein Structure and Function

Amphoteric Property of Amino Acids

• Amino acids are amphoteric, meaning they can act as both acids and bases
• This duality arises from the presence of both a carboxyl (-COOH) group and an amino (-NH2) group within their structure
• The carboxyl group can donate a proton (H+), acting as an acid, while the amino group can accept a proton, acting as a base

Peptide Formation

• The formation of peptides involves a reaction between the carboxyl group of one amino acid and the amino group of another amino acid
• This reaction, known as a dehydration reaction, releases a molecule of water and forms a peptide bond.

Essential Amino Acids

• Essential amino acids cannot be synthesized by the human body and must be obtained from the diet
• They are crucial for various bodily functions, including protein synthesis, tissue repair, and hormone production

Protein Structure: Tertiary Structure

• Tertiary structure refers to the three-dimensional arrangement of a polypeptide chain, which is crucial for the protein's overall shape and function
• This level of structure is stabilized by interactions between amino acid side chains, including hydrogen bonds, ionic bonds, disulfide bridges, and hydrophobic interactions

Protein Hydrolysis

• Protein hydrolysis is the breakdown of proteins into their constituent amino acids through the addition of water
• This process, usually catalyzed by enzymes, is essential for digestion and the recycling of amino acids within the body

Function of Protein Structure

• Protein structure dictates its function
• Specific three-dimensional shapes enable proteins to interact with other molecules, facilitating a wide range of biological activities
• Examples include enzymatic catalysis, transport of molecules, structural support, and signaling

Urea Formation: Transdeamination

• The conversion of amino groups into urea primarily involves transdeamination, a process involving the transfer of an amino group to α-ketoglutarate
• This ultimately leads to the formation of urea, the primary waste product of nitrogen metabolism

Protein Denaturation vs. Hydrolysis

• Denaturation disrupts the secondary, tertiary, and quaternary structures of a protein, altering its three-dimensional shape without breaking peptide bonds
• Hydrolysis, on the other hand, breaks peptide bonds, completely degrading the protein into its constituent amino acids

Significance of Proteins

• Proteins play critical roles in virtually every biological process, from structural support and enzymatic catalysis to transport and signaling
• They contribute to cell growth, repair, and immune function

Peptide Bond Formation

• Peptide bonds are formed between the carboxyl group of one amino acid and the amino group of another
• These bonds are responsible for linking amino acids together to form polypeptide chains, the building blocks of proteins

Early Misconception about Proteins

• Early physiologists mistakenly believed that proteins were the sole source of energy for the human body
• This misconception was later corrected with the discovery of carbohydrates as the primary energy source

Standard Amino Acids in Humans

• Standard amino acids are the 20 amino acids commonly found in proteins of all living organisms
• They include alanine (Ala), arginine (Arg), asparagine (Asn), aspartic acid (Asp), cysteine (Cys), glutamine (Gln), glutamic acid (Glu), glycine (Gly), histidine (His), isoleucine (Ile), leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), and valine (Val)

'Proteios' and Protein Nomenclature

• The term 'proteios' comes from the Greek word meaning "of primary importance," reflecting the fundamental role of proteins in biological systems
• This term was used by Jöns Jacob Berzelius in the early 19th century to describe these complex organic molecules

Characteristic Feature of Proteins

• Proteins are large, complex polymers composed of amino acids linked by peptide bonds
• They exhibit a wide variety of shapes and functions, crucial for all living organisms

Protein Content in Cell Dry Weight

• Proteins typically account for a significant portion of a cell's dry weight, often around 50%
• This highlights the abundance and importance of proteins in cellular processes

Coining of the Term 'Protein'

• Jöns Jacob Berzelius, a Swedish chemist, coined the term "protein" in 1838
• This term was chosen to reflect the essential nature of these molecules for living organisms

Amino Acid in Positively Charged Form

• In highly acidic solutions (low pH), amino acids primarily exist in their positively charged forms
• This is because the carboxyl group is protonated (-COOH)
• The amino group is also protonated (-NH3+), contributing to the overall positive charge

Zwitterion Form of an Amino Acid

• A zwitterion is a molecule with both a positive and a negative charge, but an overall neutral charge
• In amino acids, the zwitterion form exists at a specific pH, typically near the isoelectric point
• At this pH, the carboxyl group is deprotonated (-COO-) and the amino group is protonated (-NH3+), resulting in a neutral net charge

Amino Acid Behavior with pH Increase

• As the pH of a solution increases (becomes more basic), an amino acid undergoes deprotonation
• The carboxyl group loses its proton, transitioning from -COOH to -COO-
• At sufficiently high pH, the amino group may also deprotonate, transitioning from -NH3+ to -NH2

Role of OH- Ions in Amino Acid Ionization

• OH- ions, present in basic solutions, can react with the protonated carboxyl group (-COOH), removing the proton and forming water (H2O)
• This process contributes to the deprotonation of the carboxyl group, leading to the formation of the carboxylate anion (-COO-)

Amino Acid Form in Basic Solutions

• In basic solutions (high pH), amino acids primarily exist in their negatively charged forms
• This is due to the deprotonation of both the carboxyl group (-COO-) and the amino group (-NH2)

Amino Acid Representation in Neutral pH

• In a neutral pH solution, amino acids typically exist as zwitterions
• This representation shows both the carboxyl group as a negatively charged carboxylate (-COO-) and the amino group as a positively charged ammonium ion (-NH3+)

R Group of a Standard Amino Acid

• The R group (also called the side chain) represents the variable portion of an amino acid
• It is attached to the central carbon atom and determines the chemical properties of the amino acid
• R groups can be hydrophobic, hydrophilic, acidic, basic, or polar, contributing to the diversity of amino acids and their unique roles in protein structure and function

Alanine in Acidic Solution

• In an acidic solution, the carboxyl group of alanine will be protonated (-COOH) and the amino group will be protonated (-NH3+)
• This results in a positively charged alanine molecule
• Alanine will exist as a cation in acidic solutions

Aspartic Acid zwitterion form

• Aspartic acid primarily exists as a zwitterion at a pH of approximately 2.9
• This is because its isoelectric point (pI) is around 2.9

Isoelectric Point (pI)

• The isoelectric point (pI) is the pH at which an amino acid or protein carries no net electrical charge
• At the pI, the amino acid exists as a zwitterion
• The pI value is significant for separating and purifying amino acids and proteins using electrophoresis

Calculating the pI of Aspartic Acid

• The pI of aspartic acid can be calculated as the average of the two pKa values associated with the carboxyl groups
• The pKa values for aspartic acid are approximately 2.1 and 3.9
• pI = (2.1 + 3.9) / 2 = 3

Negative Ion Form of Aspartic Acid at pH above 3.9

• At pH values above 3.9, which is the pI of aspartic acid, the concentration of the negatively charged form of aspartic acid will increase
• This is because the carboxyl group loses its proton, transitioning from -COOH to -COO-

Buffer Solution in Paper Electrophoresis

• A buffer solution is used in paper electrophoresis to maintain a constant pH
• This is crucial for ensuring consistent and reliable separation of amino acids based on their charges

Separating Amino Acids using Electrophoresis

• Electrophoresis separates molecules based on their charge and size using an electric field
• The net charge of an amino acid at a specific pH determines its migration direction in the electric field
• Positively charged amino acids will migrate towards the negative electrode (cathode), while negatively charged amino acids will migrate towards the positive electrode (anode)

pKa Values for Isoelectric Point Calculation of Aspartic Acid

• The two pKa values relevant for calculating the isoelectric point (pI) of aspartic acid are:
  • The pKa value of the α-carboxyl group (approximately 2.1)
  • The pKa value of the side chain carboxyl group (approximately 3.9)

Aspartic Acid at pH 9.9

• At pH 9.9, aspartic acid will exist primarily as a negatively charged species
• This is because both the α-carboxyl group and the side chain carboxyl group will be deprotonated (-COO-)

Apoenzyme Definition

• An apoenzyme refers to an enzyme that is catalytically inactive in the absence of its cofactor
• It represents the protein component of a holoenzyme, lacking the non-protein component required for full enzymatic activity

Function of Coenzymes

• Coenzymes act as non-protein organic molecules that bind to enzymes and assist in catalyzing reactions
• They often act as carriers of electrons, functional groups, or atoms, contributing to the enzyme's catalytic activity

Oxidation-Reduction Enzymes

• Oxidoreductases are a class of enzymes specifically responsible for catalyzing oxidation-reduction reactions
• These reactions involve the transfer of electrons or hydrogen atoms between molecules

'-ase' Suffix in Enzyme Names

• The '-ase' suffix is commonly used to indicate an enzyme
• This naming convention helps distinguish enzymes from other types of proteins

Tightly Bound Coenzyme

• A prosthetic group is a type of coenzyme tightly bound to an enzyme
• It is essential for the enzyme's activity and cannot be easily removed

Hydrolase Enzyme Example

• Lipases are classified as hydrolases
• They catalyze the breakdown of lipids (fats) by adding water to break the ester bonds

Transferase Enzyme Reaction

• Transferases typically catalyze the transfer of a functional group from one molecule to another
• Examples include kinases, which transfer phosphate groups

Digestive Enzyme with Older Name

• Pepsin, a digestive enzyme responsible for protein breakdown in the stomach, is known by its older name, pepsin
• This reflects its historical significance and its role in the digestive process

Amino Acid Migration at pH 6

• At pH 6, aspartic acid and arginine will migrate towards the anode (positive electrode) due to their net positive charge
• Alanine and isoleucine will not migrate due to their neutral charge at this pH
• Lysine will migrate towards the cathode (negative electrode) because it carries a net positive charge

Reason for Alanine and Isoleucine Non-Migration

• Alanine and isoleucine do not migrate in the electrophoresis at pH 6 because their isoelectric points are close to 6
• This means they have a near-neutral charge at this pH, resulting in minimal movement in the electric field

Amino Acid Migration Towards the Cathode at pH 6

• Lysine will migrate towards the cathode (negative electrode) at pH 6 because its pI is greater than 6
• This means it will have a net positive charge at pH 6, causing it to move towards the negative electrode

Ninhydrin Solution Effect on Visualization

• Spraying ninhydrin solution after electrophoresis reacts with amino acids, producing a purple color that allows visualization
• This color reaction makes it easier to identify and analyze the separated amino acids

Amino Acid With Highest pI

• In the provided sample, lysine has the highest pI (pI = 9.7)
• This indicates that lysine will exist as a zwitterion at a more basic pH compared to the other amino acids

Aspartic Acid Ion Form at pH 6

• At pH 6, aspartic acid will exist primarily in its negatively charged form
• This is because its pI is around 2.9, and at pH 6, it will be deprotonated

Difference in Migration of Amino Acids

• At pH 6, aspartic acid will migrate towards the anode (positive electrode), while alanine, isoleucine, and lysine will exhibit different migration patterns
• Alanine and isoleucine will not migrate since their pI is close to 6
• Lysine will migrate towards the cathode (negative electrode) due to its higher pI

Amino Acid Not Migrating Towards Anode at pH 10

• At pH 10, aspartic acid, which has a pI of around 3, will likely be negatively charged and migrate towards the anode
• The other amino acids might migrate differently, depending on their individual pI values
• It is important to note that all amino acids, including aspartic acid, can be negatively charged under very basic conditions (high pH values)