lec 13-Genetics and Molecular Biology: Gene Mutations Flashcards

Introduction to Gene Mutations and Molecular Biology

• Definition of Mutation: A change in the DNA sequence.

• Prevalence: Every individual organism carries mutant alleles. These may or may not manifest as distinct, observable phenotypes.

• Occurrence: New mutations can arise in every generation.

• Gene Mutations: These occur specifically within individual genes as a consequence of changes in the nucleotide sequence. This can lead to alterations in protein structure or gene expression.

• Sources of Mutation:     - DNA replication errors during the synthesis of new strands.     - Spontaneous mutations occurring without external influence.     - Chemical agents and irradiation exposure (e.g., ultraviolet light).

Classification of Biological Mutations

• Somatic Mutations:     - Occur in any cell of the body except for the germ cells (egg and sperm).     - These mutations are not inherited by offspring.     - Somatic mutations are a significant factor in the development of cancer.

• Germ-line Mutations:     - Occur in the gametes.     - These mutations are inherited by offspring and can be passed through generations.

Base-Pair Substitution Mutations

• Definition: The replacement of one nucleotide base pair with another.

• Three Primary Types:     1. Silent Mutation (Synonymous Mutation): A base-pair change that does not result in a change to the amino acid sequence. This is possible due to the redundancy of the genetic code, where multiple codons specify the same amino acid.     2. Missense Mutation: A base-pair change that results in the substitution of one amino acid for another in the resulting protein.     3. Nonsense Mutation: A base-pair change that converts a codon specifying an amino acid into a translation termination (stop) codon, leading to premature termination of translation.

Detailed Analysis of Missense Mutations

• Variations in Severity: The impact of a missense mutation depends on whether the substitution is conservative or non-conservative.

• Conservative Amino Acid Substitution:     - Substitutes one amino acid for another that is chemically similar.     - Less likely to significantly alter the protein's overall function.     - Often referred to as a neutral mutation.     - Example: Substituting Alanine ($-CH_3$ side chain) with Glycine ($-H$ side chain).

• Non-conservative Amino Acid Substitution:     - Substitutes one amino acid for another that is chemically different.     - More likely to alter the protein's function.     - The consequences depend on the role of that specific amino acid within the protein's 3D structure.     - Example: Substituting Glycine ($-H$ side chain) with Glutamate ($-CH_2-CH_2-COO^-$ side chain).

The Genetic Code and Mutation Sensitivity

• Redundancy: Many amino acids are encoded by more than one codon.

• Position Sensitivity: A mutation occurring in the first or second letter of a three-letter codon is significantly more likely to produce a change in the amino acid than a mutation in the third letter (the "wobble" position).

Consequences of Nonsense and Frameshift Mutations

• Nonsense Mutations: Result in the premature termination of translation. The produced polypeptides are truncated and are typically nonfunctional.

• Frameshift Mutations: Caused by the addition or deletion of one or more base pairs (not in multiples of three). This shifts the reading frame and alters every amino acid sequence downstream of the mutation site.

Regulatory Region Mutations

• Concept: Some point mutations do not change the amino acid sequence of the protein but instead alter the amount of protein produced.

• Targets: These mutations affect regulatory regions such as promoters and other protein-binding sites.

• Promoter Mutations:     - These interfere with the efficiency of transcription initiation.     - Impact the binding of RNA polymerase or regulatory proteins.     - Effects: Can cause mild to moderate reductions in transcription, completely abolish transcription, or in some cases, enhance transcription.     - Contextual Example: In the lac operon, mutations can occur in the RNA polymerase binding site or the repressor protein binding site.

Spontaneous Mutations and Replication Errors

• Induced Mutations: Caused by mutagens, which are environmental agents that alter nucleotide sequences through a process called mutagenesis.

• Spontaneous Mutations: Arise in the absence of known mutagens, often due to errors in DNA replication. They provide the "background rate" of mutation.

• DNA Polymerase Proofreading: DNA polymerase can mistakenly insert the wrong nucleotide. This is corrected by the $3' \rightarrow 5'$ exonuclease function of the DNA polymerase approximately $99\%$ of the time.

• Trinucleotide Repeat Disorders:     - Caused by DNA polymerase "slipping" during replication, which increases the number of trinucleotide repeats within a gene.     - This results in longer stretches of the same amino acid in the protein.     - Example: Huntington’s Disease: An autosomal dominant disorder caused by an increase in the length of a polyglutamine region in the Huntingtin protein.         - Normal: $28$ CAG repeats.         - Disease symptoms: $36$ or more CAG repeats.

Spontaneous Nucleotide Base Changes (Tautomeric Shifts)

• Tautomers: DNA nucleotide bases can convert into alternative structures called tautomers, differing in hydrogen placement and bonding.

• Tautomeric Forms:     - Adenine and Cytosine: Normal form is Amino ($C-NH_2$); Rare form is Imino ($C=NH$).     - Thymine and Guanine: Normal form is Keto ($C=O$); Rare form is Enol ($C-OH$).

• Mechanism: Tautomeric shifts can lead to base-pair mismatches (e.g., rare enol form of Guanine pairing with Thymine). If these mismatches are not repaired before the next round of replication, they result in permanent mutations in the progeny DNA.

Transposable Elements (Transposons)

• Overview: Found in the genomes of all prokaryotes and eukaryotes.

• Types:     1. DNA Transposons: Found in both prokaryotes and eukaryotes. They move via the protein transposase.     2. Retrotransposons: Found in eukaryotes. They move via an RNA intermediate and require reverse transcriptase and integrase.

• Insertion Mutations: Transposons can cause mutations by inserting into a gene, often resulting in premature translation termination and a nonfunctional protein product.

• Bacterial IS Elements (Insertion Sequences):     - Found in E. coli and the F-plasmid.     - Approximately $1000\,bp$ long.     - Contain a transposase gene bracketed by short Inverted Repeat (IR) sequences.     - Mechanism: Transposase binds to the IRs, makes blunt end cuts on the IS element, and staggered cuts at a new target site in the genome.

Types of Induced Mutations and Chemical Mutagens

• Mutagens: Physical, chemical, or biological agents.

• Specific Chemical Agents:     - Nucleotide Base Analogs: Similar to nitrogenous bases but have altered pairing properties. Example: $5$-bromouracil ($5$-BU) replaces thymine and causes an $AT \rightarrow GC$ transition.     - Alkylating Agents: Add methyl ($-CH_3$) or ethyl ($-CH_2-CH_3$) groups to bases. Example: Ethyl methanesulfonate (EMS).     - Deaminating Agents: Remove the amino ($-NH_2$) group from bases.     - Hydroxylating Agents: Add a hydroxyl ($-OH$) group.     - Oxidative Agents: Add oxygen or remove hydrogen.     - Intercalating Agents: Flat, planar molecules that wedge between base pairs, distorting the helix. This leads to additions or deletions of nucleotides, causing frameshift mutations. Examples: Proflavin, acridine orange, and ethidium bromide. (Note: GelRed is a related non-mutagenic agent because it cannot cross living cell membranes).

• Radiation: Ultraviolet (UV) light, X-rays, gamma rays, and cosmic rays.

DNA Repair Mechanisms

• 1. Photoreactivation Repair:     - Repairs damage such as thymine dimers (commonly caused by UV light).     - Utilizes the Photoreactivation Enzyme (PRE), also known as photolyase in E. coli.     - The enzyme cleaves the bonds of the thymine dimer.     - Note: This specific mechanism is not present in humans.

• 2. Base Excision Repair (BER):     - Involves specific enzymes including DNA glycosylase (e.g., uracil DNA glycosylase), AP endonuclease, DNA polymerase, and DNA ligase.     - The damaged or noncomplementary base (e.g., Uracil appearing in DNA) is identified, excised, and replaced with the correct base (e.g., Cytosine).

• 3. Nucleotide Excision Repair (NER):     - Repairs lesions that distort the DNA helix, such as thymine dimers.     - In E. coli, the uvrA, uvrB, and uvrC proteins recognize the lesion and excise a sequence of $13$ nucleotides.     - DNA Polymerase I synthesizes the replacement strand, and DNA ligase seals the single-stranded nick.

Human Diseases Related to DNA Repair Mutations

• Xeroderma Pigmentosum:     - Individuals are typically homozygous recessive for a faulty repair gene.     - Characterized by extreme photosensitivity; exposure to light leads to freckles, intense pigment patches, and malignant warty growths (often fatal).     - Caused by mutant alleles in six different genes whose proteins function in the human nucleotide excision repair mechanism.

• Hereditary Cancer:     - Mutations in DNA repair genes significantly increase cancer risk.     - BRCA2: Linked to hereditary breast cancer.     - MSH2 and MLH1: Linked to hereditary colorectal cancer.

• Somatic Impact: Somatic mutations in DNA repair genes also contribute to the development of various cancers throughout an individual's life.