Biochemistry: Proteins & Amino Acids PDF

# **AWAEL PLATFORM** - **Proteins** are the most abundant and functionally diverse molecules in living systems. - **Proteins** are macromolecules an organic nitrogenous compound of high molecular weight, consisting largely or entirely of **a-amino acids** united together by **peptide linkages** or...

# **AWAEL PLATFORM** - **Proteins** are the most abundant and functionally diverse molecules in living systems. - **Proteins** are macromolecules an organic nitrogenous compound of high molecular weight, consisting largely or entirely of **a-amino acids** united together by **peptide linkages** or **amide bond 'covalent bond'**. - **Proteins** may be **structural**, **regulatory**, **contractile**, or **protective**; they may serve in **transport**, **storage**, or **membranes**; or they may be **toxins** or **enzymes**. - **Elemental composition of proteins:** - Proteins are predominantly constituted by five major elements in the following proportion. - **Carbon:** 50-55% - **Hydrogen:** 6-7.3% - **Oxygen:** 19-24% - **Nitrogen:** 13-19% - **Sulfur:** 0-4% # **Amino acids:** - **Amino acids** are the monomeric units or building blocks of proteins. - Although more than 300 different amino acids have been described in nature, only 20 are commonly found as constituents of mammalian (Human) proteins. These are the only amino acids that are coded for by DNA, the genetic material in the cell. - Some proteins contain additional amino acids that arise by the post-translational modification of an amino acid already present in a peptide. - **Nomenclature**: By the first three letters of their name. e.g. Serine (ser) - **Exception**: Tryptophan (Trp), Isoleucine (Ile) Asparagine (Asn) & Glutamine (Gln) # **Alpha (α)- Amino acids** - The major building block of proteins are called alpha (a) amino acids. - **Amino acids** are defined as a molecules containing an amine group (-NH2), carboxyl group (-COOH), and a distinctive side chain ("R-group") bonded to the **a-carbon atom**. The overall amino acid formula can be represented as: - $R-CH(NH_2)-COOH$ - All the standard amino acids except **proline** have both free **a-amino** and free **a-carboxyl groups**. **Proline** is has a cyclic structure and contains a **secondary amine group** (called an **imino group**) instead of a **primary amine group** (called an **amino group**). - **Proline**: It differs from other amino acids in that proline's side chain and **a-amino N** form a **rigid**, **five-membered ring structure**. - Proline, then, has a **secondary** (rather than a **primary**) **amino group**. - It is frequently referred to as an **imino acid**. # **Stereochemistry of a-amino acids** - **Optical isomers** differ from each other in their **optical activity** of rotate the plane of **polarized light** either to the left or to the right. The term **levorotatory** (L or -) is used for the substances which rotate the plane of polarized light to the left. On the other hand, the term **dextrorotatory** (D or +) is used for substances rotate the plane of polarized light to right. - **Optical isomers** or **enantiomers** occur due to the presence of an **asymmetric carbon** (a **chiral carbon**; a carbon atom is attached to four different groups). The **amino acids** (except **glycine**) possess four distinct groups (R, H, COO, NH3+) held by **a-carbon**. Thus all the **amino acids** (except **glycine** where R= H) have **optical isomers**. - **Enantiomers** (D- and L-isomers being mirror images of each other), D and L enantiomers rotate polarized light in equal, but opposite directions. - **Racemic mixture** is inactive and optical rotation zero "when equal amounts of dextrorotatory and levorotatory isomers are present (50% D: 50% L)". - Most biological amino acids have the **L-configuration**. # **Amino Acids Classification** - Amino acids can be classified by different classification: - **1. Chemical classification.** - **2. Nutritional classification.** - **3. Metabolic classification.** ## 1. **CHEMICAL CLASSIFICATION** - Classification based on the **solubility** (i.e., **polarity** and **ionization**) of the side chains (**R-groups**): - **1. Hydrophilic:** 11AAs: **Arginine**, **Asparagine**, **Aspartic acid**, **Cysteine**, **Glutamic acid**, **Glutamine**, **Glycine**, **Histidine**, **Lysine**, **Serine**, and **Threonine**. - **2. Hydrophobic:** 9AAs: **alanine**, **phenylalanine**, **valine**, **leucine**, **isoleucine**, **tryptophan**, **tyrosine**, **methionine**, and **proline**. ## **TABLE: Hydrophilic & Hydrophobic Amino Acids** | Hydrophilic | Hydrophobic | |----------------|------------------| | Arginine | Alanine | | Asparagine | Isoleucine | | Aspartic acid | Leucine | | Cysteine | Methionine | | Glutamic acid | Phenylalanine | | Glutamine | Proline | | Glycine | Tryptophan | | Histidine | Tyrosine | | Lysine | Valine | | Serine | | | Threonine | | ## **1. Chemical Classification** - Classification based on the **polarity** of the side chains (**R-groups**): - **1. Nonpolar:** 9AAs: **alanine**, **phenylalanine**, **valine**, **leucine**, **isoleucine**, **tryptophan**, **methionine**, and **proline**. - **2. Polar (charged & non-charged):** These amino acids may be subclassified into: - **a. Polar negatively charged (acidic)** - 2AAs: Both **aspartic** and **glutamic acids** are acidic amino acids. They are proton donors. At physiologic pH, the side chains of these amino acids are fully ionized, containing a negatively charged carboxylate group (-COO¯). They are, therefore, called **aspartate** or **glutamate**. - **b. Polar positively charged (basic)** - 3AAs: **histidine**, **lysine**, and **arginine** are Basic amino acids. The side chains of the basic amino acids accept protons. At physiologic pH the side chains of **lysine** and **arginine** are fully ionized and positively charged. In contrast, **histidine** is weakly basic, and the free amino acid is largely uncharged at physiologic pH. However, when histidine is incorporated into a protein, its side chain can be either positively charged or neutral, depending on the ionic environment provided by the polypeptide chains of the protein. - **c. Polar Neutral (unionized or non-charged)** - 6AAs: **Serine**, **threonine**, **tyrosine**, **asparagine**, **glutamine**, and **cysteine**. ## **2. Nutritional classification** - They are classified into three groups: - **Essential (9 aa):** They are amino acids which **cannot** be synthesized in the body and **must** be taken in diet; **Histidine**, **Methionine**, **Threonine**, **Valine**, **Leucine**, **Isoleucine**, **Lysine**, **Phenylalanine**, and **Tryptophan**. - **Non essential (11 aa):** can **be** synthesized inside the body as **Alanine**, **Arginine**, **Asparagine**, **Aspartate**, **Glutamine**, **Glutamate**, **Glycine**, **Proline**, **Serine**, **Cysteine**, and **Tyrosine**. - **Semi essential aas:** Some aas can become **conditionally essential**. For example, supplementation with **glutamine** and **arginine** has been shown to improve outcomes in patients with trauma, post-operative infections, and immunosuppression. **Arginine & Histidine** can be also classified as: Semi essential amino acids, as they are formed in the body in amount enough for adults, but not in during growth of children or during pregnancy and lactation. ## **TABLE: Essential, Non-Essential, Glucogenic, Ketogenic amino acids** | Type | Essential | Nonessential | Glucogenic | Ketogenic | |---------------------|-------------------|-----------------------|-------------------|--------------------| | Essential | Histidine | Alanine | Alanine | | | | Methionine | Arginine | Arginine | | | | Threonine | Asparagine | Asparagine | | | | Valine | Aspartate | Aspartate | | | | | Cysteine | Cysteine | | | | | Glutamate | Glutamate | | | | | Glutamine | Glutamine | | | | | Glycine | Glycine | | | | | Proline | Proline | | | | | Serine | Serine | | | | Isoleucine | | Tyrosine | | | | Phenylalanine | | | Isoleucine | | | Tryptophan | | | Leucine | | | | | | Lysine | | Non-Essential | | | | | ## **3. Metabolic classification** According to their fate in the body they are classified into three groups: - **Glucogenic amino acids (14 aas):** give glucose inside the body as **Alanine**, **Arginine**, **Asparagine**, **Aspartate**, **Cysteine**, **Glutamate**, **Glutamine**, **Glycine**, **Proline**, **Serine**, **Histidine**, **Methionine**, **Threonine**, and **Valine**. - **Ketogenic amino acids (2 aas):** gives ketone bodies as **Leucine** and **Lysine**. - **Glucogenic and ketogenic amino acids (4 aas):** They give both glucose and ketone bodies as **Tyrosine**, **Isoleucine**, **Phenylalanine**, and **Tryptophan**. # **Properties of Amino Acids:** ## **1. Physical Properties of Amino Acid Solutions** - AAs are **colorless**, **crystalline** substances. Most AAs are **tasteless** but some are sweet (e.g. Glycine, Alanine) and some are bitter (e.g. Arginine). - **SOLUBILITY:** Solubility of aas depends upon **polarity** of the **solvent**, **iso-electric point**, **pH**, and **temperature**. - **Polarity of the solvent:** High solubility in polar solvents (i.e. water and ethanol), and low solubility in non-polar solvents (i.e. benzene, chloroform, ether). - **Isoelectric point:** AAs are insoluble at an iso-electric point (pl) due to net neutral charge. At the isoelectric point, where the net charge on the amino acid is zero. - **Temperature** - Solubility can increase with higher temperatures "Tyrosine is soluble in hot water". - The **MELTING POINT** of aas is high due to the strong ionic attractions within the **zwitterionic structure**. - Amino acids exist as **zwitterions**, where the carboxyl group is deprotonated and the amino group is protonated, leading to strong electrostatic attractions between the charged groups. These strong ionic interactions result in the amino acids being crystalline solids with high melting points, often in the range of 200°C to 300°C. ## **Chemical Properties of Amino Acid Solutions** - **Acidic and Basic of Amino Acids** - **Acts as buffer:** AA can react with both bases (OH-) and acids (H+) to form salts (conjugate base or conjugate acids). - **In acid solution**, aas carry **positive charges** and hence they move towards **cathode** in an electric field. - **In an alkaline solution**, the AAs carry **negative charges** and therefore move towards the **anode**. - **In an water**, it exists as inner salt carrying both **positive** and **negative charges**. This occurs as a result of the dissociation of the carboxyl group to release the Ht ion, which passes from the carboxyl to the amino group. - **Amphoteric property:** Amino acids are amphoteric, meaning they can act as both acids and bases due to their zwitterionic structure. When an amino acid contains both positive charge and a negative charge in the "backbone", it is called a **zwitterion** and has an overall neutral charge. - The zwitterion of an amino acid exists at a pH equal to the **isoelectric point**. - The **isoelectric point (pl)** is the pH at which the net charge on the amino acid molecule is zero, as the sum of the positive and negative charges within the molecule is equal. - The pl is a characteristic property of each individual amino acid, determined by the pka values of the functional groups (amine and carboxyl) in the molecule. - In aqueous solutions, amino acids can exist in different protonation states, gaining or losing protons at their amine and carboxyl groups, depending on the solution pH. - At **physiologic pH** (approximately pH 7.4): The carboxyl group is dissociated, forming the negatively charged carboxylate ion (-COO-), and the amino group is protonated (-NH3+). ## **Titration Curve of an Alanine** - The **isoelectric point (pI)** is the pH at which the number of positive and negative charges on a population of molecules is equal (i.e. no net charge). - The **pI of alanine** is **6.02** (calculated as (pK1+ pK2)/2) - This means that at pH 6.02, the amino group is protonated and the carboxyl group is deprotonated, resulting in a net charge of 0. - The titration curve of alanine shows the change in pH of a solution of alanine as a base (OH-) is added. ## **Titration Curve of an Aspartic Acid** - The **pI of aspartic acid** is **3.02** (calculated as (pK1 + pK3)/2) - This means that at pH 3.02, both the carboxyl groups are deprotonated and the amino group is protonated, resulting in a net charge of 0. - The titration curve of aspartic acid shows the change in pH of a solution of aspartic acid as a base (OH-) is added. ## **Titration of an Alanine** - **Buffer pairs:** The - COOH/- COO pair can serve as a buffer in the pH region around pK1, and the - NH3+/- NH2 pair can buffer in the region around pK2. - **When pH = pK:** When the pH is equal to **pK1** (2.3), equal amounts of **Forms I** and **II** of alanine exist in solution. - When the pH is equal to pK2 (9.1), equal amounts of Forms II and III are present in solution. - **Isoelectric point:** The **pI of alanine** is 6.02. It is calculated by: (pK1 + pK2)/2 ## **Titration Curve of a Histidine** - The **pI of histidine** is **7.585** (calculated as (6.0 + 9.17)/2). # **Peptides and polypeptides:** - **Act of Amino Acids** as building block of **peptides**, **polypeptides** & **proteins**. - **Amino Acid Sequence:** An amino acid sequence refers to the linear arrangement of amino acids in a polypeptide or protein molecule. - **Dipeptide:** consisting of two amino acids linked by a single peptide bond. - **Tripeptide:** consisting of three amino acids linked by two peptide bonds. - **Oligopeptides:** contain 4 to 10 amino acids. - **Polypeptides:** contain more than 10 amino acids. - **Proteins:** both in the body and diet, are large polypeptide molecules with more than 50 amino acids. ## **Formation of peptide bond and polypeptide chain** - **The Peptide Bond:** In proteins, amino acids are joined covalently by **peptide bonds**, which are **amide linkages** between the **a-carboxyl group** of one amino acid and the **a-amino group** of another. The reaction is a dehydration (elimination of a molecule of water). - **Since all amino acids contain both NH2 and COOH groups**, long chains of amino acids may be formed. - **The resultant chain of amino acids therefore has an amino group at one end (the N-terminus) and a carboxyl group at the other end (the C-terminus).** ## **Characteristics of the peptide bond:** - **1. Resonance properties** - A peptide bond is a **planar**, **trans**, and **rigid configuration**. It also shows a **partial double bond character**. - The **coplanarity** of the peptide bond denotes the **resonance** or **partial sharing of two pairs of electrons** between the **amide nitrogen** and **carboxyl oxygen**. - **Because of this resonance**, The peptide bond is shorter than a single bond, and is rigid and planar. This prevents free rotation around the bond between the carbonyl carbon and the nitrogen of the peptide bond. However, the bonds between the a-carbons and the a-amino or a-carboxyl groups can be freely rotated (although they are limited by the size and character of the R-groups). - A **trans configuration** is more stable than a **cis configuration**, because a steric clash between the side chains. - **2. Resonance properties & Trans configuration** - The peptide bond has partial double bond character. - The peptide bond has **resonance structures (shown above)**, due to partial sharing of two pairs of electrons. - Therefore: - **C-N bond** is shorter than C-N bond of a simple amine; - **O has 8- charge, N has 8+ charge** (there in an electric dipole); - **peptide bond does not rotate**. - **3. Polarity of the peptide bond** - Like all amide linkages, the peptide bond is uncharged, and neither accept nor release protons over the pH range of 2-12. Thus, the charged groups present in polypeptides consist solely of the N-terminal (a-amino) group, the C-terminal (a- carboxyl) group, and any ionized groups present in the side chains of the constituent amino acids. - The -C=O and -NH groups of the peptide bond is polar, and are involved in hydrogen bonds. ## **Unusual peptide (Glutathione)** - Glutathione is a tripeptide with the sequence **y-glutamylcysteinylglycine**. - It has a unique structure, in which the carboxyl group of glutamic acid is linked to the amino group of cysteine through a gamma (y) linkage. This gives glutathione a unique functionality and it is involved in various important cellular processes. - **Reduced glutathione** has a **thiol group (-SH)** in its cysteine residue. The thiol group allows glutathione to act as a **reducing agent** and it help protect cells from oxidative damage. - **Oxidized glutathione** is formed when two molecules of reduced glutathione react, forming a disulfide bond (-S-S-) between their cysteine residues.