Genetic Mutations and Protein Sequences

• Overview of how mutations in the genetic code affect protein sequences
• The lesson dives into DNA sequences and mutations and how they alter protein production.

DNA to RNA to Protein

• DNA is transcribed into messenger RNA (mRNA)
• mRNA carries the genetic code to the ribosome for translation
• The top DNA strand is the template for RNA synthesis
• RNA matches the DNA sequence of the top strand, with thymine (T) replaced by uracil (U)
• Codons (three nucleotide sequences) in mRNA translate into amino acids, forming proteins
• Charts show the amino acid sequence based on RNA or DNA, no need to memorize
• Example amino acid sequence: Tyrosine (Tyr), Glycine (Gly), Leucine (Leu), Leucine (Leu)

Point Mutations

• Point mutations alter a single base pair in the DNA sequence
• Can change a protein's amino acid sequence

Three Types of Point Mutations

• Missense Mutation: changes a single DNA base, resulting in a different amino acid
• Example: Sanger sequencing trace shows a mutation from glycine (Gly) to aspartic acid (Asp)
• Impact: Varies depending on the amino acid's role in protein function
• Synonymous Mutation: changes a single DNA base, but the same amino acid is encoded due to codon redundancy
• Example: A mutation in the third guanine of the codon, but the protein sequence remains unchanged
• Impact: Usually benign, but can have subtle effects depending on gene expression
• Nonsense Mutation:changes a single DNA base, creating a stop codon within the coding sequence
• Example: A mutation in the DNA creates a TAA stop codon early in the protein sequence
• Impact: This results in a truncated, often nonfunctional protein

Insertions and Deletions (Indels)

• Insertion mutations add extra bases into the DNA sequence
• Deletion mutations remove bases from the DNA sequence

Frame-shift Insertion

• Adding an extra base into the sequence shifts the reading frame causing an incorrect amino acid sequence
• Example: Adding a base after the first two guanines
• Impact: If not at the very end of the sequence, severe disruption of protein function will occur

Frame-shift Deletion

• Deleting a single base shifts the reading frame, changing the amino acid sequence
• Example: Deleting a guanine will cause the protein to produce incorrect amino acids after the deletion
• Impact: Similar to insertions, this can lead to a nonfunctional or incomplete protein

In-Frame Mutations

• Inserting or deleting exact multiples of three bases in the sequence maintains the reading frame
• Mutation does not shift the entire sequence
• Example: Inserting or deleting three bases
• Impact: Less damaging than frame-shifting mutations but may still lead to functional changes

Splice Mutations

• Occur at the boundaries between exons and introns and can disrupt gene splicing during RNA processing
• Single base substitutions or larger insertions/deletions affecting the splicing process
• Cause exons to be skipped or create incorrect exon-intron boundaries and this may alter the final protein
• Impact: Can lead to missing or incorrect protein segments, potentially affecting protein function

Summary of Mutation Types

• Single Base Substitutions: Missense, synonymous, nonsense, and splice site mutations
• Insertions and Deletions: In-frame insertions/deletions, frame-shift insertions/deletions

Impact of Mutations on Protein Function

• Frame-shift mutations: Likely to cause severe damage to the protein, especially if near the beginning of the coding sequence
• Nonsense mutations: Likely to result in a nonfunctional protein
• Splice site mutations: Can cause significant changes to the protein if they cause exons to be skipped or incorrect regions to be included
• In-frame mutations: Generally less damaging, but in rare cases can disrupt protein function
• Synonymous mutations: Often benign, but can affect transcription or splicing due to altered regulatory elements
• Missense mutations: Impact can be variable, with some leading to disease and others having minimal effects

Functional Importance of Specific Proteins

• Duplicated Genes: Some genes are so vital that they have been duplicated in the genome
• Example: rRNA in the genome, losing one copy typically doesn't have a major impact
• Other genes, such as olfactory receptors, have multiple versions but are not survival-critical
• Vital Genes with Single Copies: Some genes such as those encoding for fibrillin-1 (associated with Marfan Syndrome) are essential for survival
• Mutations in these genes are more likely to result in disease
• Example: A missense mutation in fibrillin-1 can lead to serious diseases like Marfan syndrome

Intergenic and Deep Intronic Mutations

• Mutations outside of protein-coding regions (intergenic mutations) or deep within introns (deep intronic mutations) may not directly alter the protein sequence
• ENCODE Project (2012): Showed that 76% of the human genome is transcribed, and over 80% is involved in some form of DNA-protein interaction
• Regions play a crucial role in regulating gene expression and transcription rates, indirectly affecting protein levels in cells

Regulation of Transcription

• Mutations that affect regulatory regions can alter transcription, influencing how much of a protein is made
• Even intergenic or intronic mutations can affect the regulation of gene expression and protein function

Assessing Mutations

• Considering the role of the ENCODE project's findings, mutations in non-coding regions can affect gene regulation and transcription levels, potentially impacting protein levels
• Understanding that a large percentage of the human genome is transcribed influences the assessment of mutations in non-coding regions revealing that they may affect gene expression, regulation, and transcription levels, thereby impacting protein levels

Synonymous Mutation Impact

• Synonymous mutations can influence mRNA structure and stability, thereby affecting translation efficiency, which can be especially relevant in genes with multiple isoforms
• Synonymous mutations, where the amino acid stays the same, can affect mRNA structure or stability through codon usage
• Certain codons are used more often than others in the genome, which is known as codon usage bias
• If a synonymous mutation changes the codon from one version to another, it may affect the speed at which the ribosome translates that codon
• Slower translation or disrupted translation timing can affect the folding of the protein or the production of isoforms if alternative splicing is involved

mRNA Secondary Structure

• mRNA molecules can fold into secondary structures, important for the stability of the mRNA
• Structures can include hairpins, loops, or stem-loops that form when specific sequences of nucleotides come together
• Translation is affected by these folds regarding how easily the ribosome can access the mRNA to translate it
• Synonymous mutation can change the sequence of the mRNA, potentially causing a different secondary structure
• For instance, the change from one codon to another might cause the mRNA to fold in a way that makes it harder for the ribosome to bind or may stabilize certain mRNA structures that could delay translation or alter splicing patterns