DNA, RNA, and Protein Synthesis

DNA to RNA to Protein

• DNA transcribes into RNA, specifically mRNA, which carries the genetic code
• The top DNA strand is the template for RNA synthesis
• RNA matches the DNA sequence of the top strand, except thymine (T) is replaced with uracil (U)
• Codons (three nucleotide sequences) in mRNA translate into corresponding amino acids, forming a protein
• Charts can be used to look up the amino acid sequence based on the RNA or DNA sequence
• Example sequence: Tyrosine (Tyr), Glycine (Gly), Leucine (Leu), Leucine (Leu)

Point Mutations

• Point mutations affect a single base pair in the DNA sequence, potentially altering the amino acid sequence of a protein
• Three types of point mutations exist: missense, synonymous, and nonsense

Missense Mutation

• A single DNA base changes, resulting in a different amino acid in the protein sequence
• A Sanger sequencing trace shows a mutation from glycine (Gly) to aspartic acid (Asp)
• The impact varies depending on the amino acid's role in protein function

Synonymous Mutation

• A single DNA base changes, but the same amino acid is still encoded
• Mutation occurs in a redundant codon
• The third guanine mutation still results in the same amino acid, and the protein sequence remains unchanged
• Usually benign, but can have subtle effects depending on the context of the gene expression

Nonsense Mutation

• A single DNA base change leads to the creation of a stop codon within the coding sequence
• A mutation in the DNA creates a TAA stop codon early in the protein sequence
• Results in a truncated protein that is often nonfunctional

Insertions

• Insertion mutations involve adding extra bases into the DNA sequence
• Frame-shift Insertion: An extra base is inserted into the sequence, shifting the reading frame, causing an incorrect amino acid sequence from that point onward
• Adding a base after the first two guanines shifts the reading frame, changing the amino acid sequence entirely
• Causes severe disruption of protein function in it is not at the very end of the sequence

Deletions

• Deletion mutations involve removing bases from the DNA sequence
• Frame-shift Deletion: If a single base is deleted, the reading frame is shifted again, changing the amino acid sequence significantly
• Deleting a guanine will cause the protein to produce incorrect amino acids after the deletion
• Similar to insertions, this can lead to a nonfunctional or incomplete protein

In-Frame Mutations

• In-frame Insertions or Deletions: Inserting or deleting exact multiples of three bases in the sequence, maintains the reading frame
• Inserting or deleting three bases results in a minor change to one or two codons but does not affect the rest of the protein
• Less damaging than frame-shifting mutations but may still lead to functional changes

Splice Mutations

• Occur at the boundaries between exons and introns, disrupting RNA processing
• The mutations can be single base substitutions, or larger insertions or deletions affecting the splicing process
• Splice mutations can cause entire exons to be skipped or incorrect exon-intron boundaries to be created, altering the final protein
• Leads 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 cause severe damage to the protein, near the beginning of the coding sequence
• Nonsense mutations: Likely to result in a nonfunctional protein
• Splice site mutations: Can result in 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, they can disrupt protein function
• Synonymous mutations: Often benign, but in some cases, they can affect transcription or splicing due to altered regulatory elements
• Missense mutations: Variable impact, some lead to disease and others have minimal effects

Functional Importance of Specific Proteins

• Some genes are so vital that they have been duplicated in the genome
• There are many copies of rRNA in the genome, and losing one copy typically doesn't have a major impact
• Other genes, such as those for olfactory receptors, have multiple versions but are not critical for survival
• Some genes related to fibrillin-1 (associated with Marfan Syndrome), are essential for survival, having a single copy means mutations in these genes are more likely to result in disease
• A missense mutation in fibrillin-1 can lead to diseases (Marfan syndrome), while mutations in olfactory receptors generally do not result in disease

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): 76% of the human genome is transcribed, and over 80% is involved in some form of DNA-protein interaction
• While these regions may not code for proteins, they 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 how much of a protein is made
• Even intergenic or intronic mutations can have an effect on the regulation of gene expression and protein function