Liu3304_fall2023_lecture_5

Insulin

The protein insulin is a polypeptide hormone crucial for glucose metabolism, consisting of two polypeptide chains known as the A and B chains, which are linked by disulfide bonds.

Amino Acid Sequence and Homology

Insulin from various organisms has been sequenced, revealing fascinating homologies. For instance, the amino acid sequence of human insulin and duck insulin differs only by six residues. The specific amino acid substitutions are as follows:

• Human Insulin: Thr, Ser, Ile, Phe, Val, Thr

• Duck Insulin: Glu, Asn, Pro, Ala, Ala, Ser

This difference in residues impacts several functional properties of the insulin, including its receptor binding affinity and its biological potency.

pI Comparison

The question arises whether the isoelectric point (pI) of human insulin is lower or higher than that of duck insulin. Understanding this requires recognizing that the substitution of Thr with Glu results in a lower pI for human insulin, indicating that human insulin has a higher pI than duck insulin. pI is crucial as it influences the solubility and stability of the protein under physiological conditions.

Charge of Peptide Chain at pH = 8

When analyzing the peptide chain TNMFDKR at pH = 8, the charge can be determined by comparing the pH to the pKa values of the functional groups in the peptide. At low pH, the predominant form is largely protonated (acidic), while at higher pH, the form tends to be deprotonated (basic).

• The peptide exists as NH2-TNMFDKR-COOH at this pH level.

• The respective pKa values are:

  • pK1: 2 (carboxylic acid group)

  • pKa2: 9 (amino group)

  • pKa for side chains: 3.9 for acidic groups and 10.5 for basic groups.

• This results in:

  • COO- representing the deprotonated form and contributing a negative charge.

  • NH3+ representing the protonated form and contributing a positive charge.

Overall Net Charge Analysis

The overall net charge of the peptide can be calculated by combining the contributions from each of these components, providing insights into the peptide’s behavior in biological systems.

Study Plan for Midterm 1

A midterm is scheduled for September 14. To prepare:

• Complete Chapter 4 and start Chapter 5.

• Note that the topic of Protein Purification has not been covered.

• Suggested practice problems in the Textbook and Student Companion for Chapters 4 and 5, particularly specific questions should be completed.

• Key calculations involve determining the pI for various amino acids and peptides, using pK values of 2 and 9 as references.

Amino Acid Stereochemistry

Understanding chirality and optical activity is essential in biochemistry. Chiral molecules can be represented using Fischer projections, adhering to the Cahn-Ingold-Prelog rules. These projections assist in visualizing the three-dimensional configurations of amino acids.

Four Levels of Protein Structure

Protein structure is categorized into four levels:

1. Primary Structure: The linear sequence of amino acids forming the polypeptide chain.

2. Secondary Structure: Localized folding patterns within the polypeptide, including alpha-helices and beta-sheets, stabilized by hydrogen bonding.

3. Tertiary Structure: The overall three-dimensional shape of the protein, dictated by interactions between side chains.

4. Quaternary Structure: Composed of multiple polypeptide chains (subunits) that assemble into a functional protein complex.

Optical Activity

Optical activity refers to the capability of a compound to rotate plane-polarized light. Enantiomers, which are mirror images of one another, exhibit different optical activities; one may rotate light dextrorotatory (+) and the other levorotatory (-). This characteristic is crucial for drug design and interactions in biological systems.

Fischer Projections

Fischer projections are a method for depicting the configuration of chiral centers in organic compounds. In the case of amino acids, L-amino acids are represented with the amino group (NH3+) on the left side, allowing for clearer visual interpretation of stereochemistry.

Amino Acids with Chiral Centers

Notably, amino acids such as threonine and isoleucine contain multiple chiral centers, leading to the existence of four stereoisomers for each. Racemic mixtures, which encompass equal proportions of enantiomers, have implications in pharmacology and biochemistry.

Cahn-Ingold-Prelog System

This system provides a standardized nomenclature for establishing the absolute configuration of chiral centers in molecules. The precedence of groups attached to the chiral center is determined by atomic mass, designating the configuration as R (rectus) or S (sinistrous).

Post-Translational Modifications (PTMs) of Amino Acids

Post-translational modifications refer to various chemical modifications that occur to amino acids after they are incorporated into a protein. Common PTMs include hydroxylation, methylation, acetylation, and phosphorylation, all of which have significant impacts on protein function, especially in cellular signaling pathways and regulatory mechanisms.

Molecules Derived from Amino Acids

Amino acids serve as the precursors for several critical bioactive molecules. For instance, GABA is derived from glutamate, dopamine from tyrosine, histamine from histidine, and thyroid hormones also originate from tyrosine. It is notable that there are approximately 250 different amino acids that have been identified in various plants and fungi, showcasing the vast diversity of this essential biomolecule in nature.

Proteins: Primary Structure

The primary structure of proteins highlights polypeptide diversity and sequencing complexities. Techniques such as Edman degradation allow researchers to determine the sequence of amino acids in a protein, contributing to our understanding of evolutionary processes and protein function across species.

Levels of Protein Structure

1. 1° Primary: The unique sequence of amino acids in a polypeptide chain.

2. 2° Secondary: Formation of structures like alpha-helices and beta-sheets through local interactions.

3. 3° Tertiary: The overall 3D structure influenced by side chain interactions and environmental conditions.

4. 4° Quaternary: The assembly of multiple polypeptide chains into a functioning protein complex.

Cytochrome C

Cytochrome C exemplifies evolutionary conservation in its function across various species, underlining essential amino acid identities necessary for its role in cellular respiration. Variability in evolutionary stages rather than linear progression is crucial for understanding phylogenetics.

DNA Mutation Rate

Significantly, DNA displays a consistent mutation rate over generations. However, crucial proteins maintain stability through conservation, preventing mutations that could otherwise lead to loss of function, emphasizing the balance between adaptability and functionality in biological systems.