Mutations and More

🧬 1. What Are Mutations?

✔ Mutations = Permanent changes in the DNA sequence.
✔ They happen naturally (random errors in replication) or due to external factors (radiation, chemicals, viruses).
✔ Most mutations are neutral, but some cause diseases or create beneficial traits (evolution!).

📌 Where Can Mutations Occur? ✔ Coding regions (exons) → Can change proteins.
✔ Non-coding regions (introns, regulatory sequences) → Can affect gene expression.

🔬 2. Types of Mutations & Which Bases Get Switched

📌 Point Mutations (Single Base Changes)

✔ A single nucleotide is swapped for another.
✔ These happen most often during DNA replication when DNA polymerase makes an error.
✔ Most common type of mutation!

📌 Types of Point Mutations:

📌 Why Do Some Base Changes Matter More Than Others?
✔ Pyrimidine-to-pyrimidine (T ↔ C) or purine-to-purine (A ↔ G) mutations are called "Transitions" → Less disruptive.
✔ Pyrimidine ↔ Purine (C ↔ A, G ↔ T, etc.) mutations are called "Transversions" → More disruptive because they change DNA helix structure.

Example of Missense Mutation: Sickle Cell Disease

✔ Normal: GAG (Glutamic Acid, polar)
✔ Mutated: GUG (Valine, non-polar)
✔ Effect: Hemoglobin sticks together, changing red blood cell shape → leads to sickle cell anemia.

🔬 3. Frame-Shift Mutations (Big Structural Changes)

✔ Caused by Insertions or Deletions (Indels) that shift the reading frame.
✔ Since codons are read in triplets, adding/removing a base shifts everything downstream, making a completely different protein.

📌 Types of Frame-Shift Mutations:

✔ Example: Tay-Sachs Disease

• A 4-base pair insertion in HEXA gene disrupts enzyme function, leading to neurodegeneration.

📌 Why Are Frame-Shift Mutations More Severe?
✔ They alter all codons after the mutation, changing multiple amino acids.
✔ Often introduces a premature STOP codon, leading to a nonfunctional protein.

🔬 4. Mutagen-Induced Mutations

✔ Environmental factors like radiation, chemicals, and viruses can damage DNA, leading to mutations.

📌 Common Mutagens & Their Effects

✔ Example: UV Light & Thymine Dimers

• UV light fuses two adjacent thymine bases (T=T), creating a kink in the DNA.

• This blocks DNA replication and transcription, leading to errors or apoptosis (cell death).

🔬 5. How Mutations Affect Protein Function

✔ Some mutations change how proteins fold, interact, or function.
✔ Protein folding is super sensitive to even small changes in amino acids.

📌 How Polarity Affects Protein Function

✔ Example: Cystic Fibrosis (CFTR Gene)

• Deletion of a single codon (ΔF508) removes Phenylalanine (F).

• This makes the CFTR protein misfold, preventing chloride ion transport.

• Leads to thick mucus buildup in lungs and digestive system.

🔬 6. Mutation Rate & Repair Mechanisms

✔ Mutations happen all the time, but cells have repair systems to fix them.

📌 Mutation Rate in Humans:

• 2-3 mutations per cell division.

• ~100 new mutations per generation (parents → child).

📌 DNA Repair Mechanisms

✔ Example: Xeroderma Pigmentosum (XP)

• Defect in Nucleotide Excision Repair (NER).

• Leads to extreme sensitivity to UV light because cells can't repair thymine dimers.

• Patients develop skin cancers at a very young age.

1. DNA Replication & Mutations

📌 Helicase Unwinds DNA

✔ Helicase is an enzyme that unzips the double-stranded DNA into two single strands.
✔ This requires ATP because breaking hydrogen bonds between base pairs takes energy.

Analogy: Think of helicase as a zipper that opens up the DNA strands for copying.

📌 Leading vs. Lagging Strand

✔ Leading Strand → DNA polymerase adds nucleotides continuously in the 5’ → 3’ direction.
✔ Lagging Strand → Synthesized in fragments (Okazaki fragments) because DNA polymerase can only work 5' → 3'.

🔬 2. tRNA & Translation

📌 What is tRNA?

✔ tRNA (transfer RNA) is a small RNA molecule that helps assemble proteins.
✔ One end carries an amino acid.
✔ The other end has an anticodon that pairs with codons on mRNA to translate the genetic code.

Example:
If the mRNA codon is AUG, the corresponding tRNA anticodon is UAC and it will bring in the amino acid Methionine (Met).

🔬 3. DNA Mutations & Damage

📌 How Often Do Mutations Happen?

✔ 2-3 mutations per cell division.
✔ Most mutations are harmless or repaired, but some cause diseases.

📌 Types of DNA Damage

📌 Why Do Thymine Dimers Matter?
✔ UV light makes two thymines next to each other form a covalent bond (T=T), distorting the DNA structure.
✔ If not fixed, it can block replication and lead to skin cancer (e.g., melanoma).

📌 What Can Cause Mutations?

✔ Radiation (UV, X-rays).
✔ Pollution (mutagens from road fumes, industrial waste).
✔ Infectious diseases (some viruses insert their own DNA into our genome).

🔬 4. Lactose Intolerance & Gene Regulation

📌 Why Are Some People Lactose Intolerant?

✔ Lactase is an enzyme that breaks down lactose (milk sugar) into glucose + galactose.
✔ The lactase gene turns off in most adults, leading to lactose intolerance.
✔ Some people have a mutation that keeps the lactase gene on, allowing them to digest milk into adulthood.

📌 This is an example of a mutation outside the gene affecting phenotype (how traits appear).

🔬 5. Mutation Effects on Proteins

✔ Some mutations change how proteins fold and function.
✔ T → C mutations (thymine to cytosine) are the most common in humans.

📌 Nonsense Mutations = Premature Stop Codon

✔ A STOP codon appears too early, making a truncated (shortened) protein.
✔ The protein loses function, which can cause genetic diseases.

📌 Sickle Cell Disease: A Classic Example of Mutation Effects

✔ Single Nucleotide Change (T → A) in the HBB Gene.
✔ Changes codon GAG (Glutamic Acid, polar) → GUG (Valine, non-polar).
✔ This disrupts hemoglobin, making red blood cells sickle-shaped and reducing oxygen transport.

📌 Why Does This Cause Problems?
✔ Normal hemoglobin is water-soluble, but the mutated version sticks together, forming long fibers.
✔ This makes red blood cells rigid and clog blood vessels.

🔬 6. COVID-19 & RNA Mutations

✔ COVID-19 uses RNA polymerase to replicate, but RNA polymerase lacks proofreading abilities.
✔ This means it makes more mistakes, causing more mutations over time.

Example:

• The Omicron variant had more mutations than earlier versions of the virus, affecting how it binds to cells.

🔬 7. Amino Acid Polarity Changes & Mutations

✔ Mutations can change the polarity of an amino acid, affecting protein folding.

📌 Examples of Polarity Changes

📌 If a polar amino acid is replaced by a non-polar one, the protein structure may collapse.

🔬 8. CRISPR-Cas9: Gene Editing

✔ CRISPR-Cas9 is a revolutionary gene-editing tool.
✔ It uses Cas9 (a bacterial enzyme) to cut DNA at specific sites.
✔ Scientists can fix mutations or introduce new genes with this system.

📌 Example: Scientists have used CRISPR to correct sickle cell mutations in experimental treatments.

🔬 9. Paxlovid vs. Molnupiravir (COVID-19 Drugs)

📌 How Do These Drugs Work?

✔ Paxlovid (more effective) → Inhibits viral protease, preventing the virus from cutting its polyprotein into functional parts.
✔ Molnupiravir → Introduces errors in viral RNA, causing it to make defective copies.

📌 Which One is Better?

• Paxlovid is ~2x more effective than Molnupiravir.

• Molnupiravir increases mutations, but this can sometimes be unpredictable.

🔬 10. AlphaFold & Protein Structure Prediction

✔ AlphaFold is an AI model that predicts the 3D structure of proteins from amino acid sequences.
✔ This is super important for drug discovery because protein shape determines function.

📌 Computational Protein Design:

• Scientists can work backward from a 3D protein shape to predict the amino acid sequence needed.

• This is useful for designing new drugs and understanding genetic diseases.

🔥 Summary Table: Biomacromolecules

1⃣ Overview: What is Transcription?

✔ Transcription = DNA → RNA (first step in gene expression).
✔ Happens in the nucleus (eukaryotes) & cytoplasm (prokaryotes).
✔ RNA polymerase reads the DNA template strand (3' → 5') and synthesizes RNA (5' → 3').
✔ Produces messenger RNA (mRNA), which carries the genetic code to ribosomes for translation.

2⃣ Stages of Transcription

📌 Transcription has three main stages:

1⃣ Initiation → RNA polymerase binds to DNA and begins synthesis.
2⃣ Elongation → RNA polymerase extends the RNA strand.
3⃣ Termination → RNA polymerase stops transcription and releases RNA.

🔬 3⃣ Step 1: Initiation of Transcription

Goal: RNA polymerase binds to a promoter region and starts making RNA.

📌 Key Players in Transcription Initiation:
✔ Promoter Region → A sequence of DNA that signals where transcription should start.
✔ RNA Polymerase → The main enzyme that synthesizes RNA.
✔ Transcription Factors (Eukaryotes only) → Help RNA polymerase bind to the promoter.

🔹 Prokaryotic Transcription Initiation (Bacteria)

✔ RNA Polymerase Holoenzyme binds directly to the promoter.
✔ Promoter contains two key regions:

• -35 Box (TTGACA) → Helps RNA polymerase recognize the promoter.

• -10 Box (TATAAT, aka Pribnow Box) → Helps DNA unwind for transcription to start.

🔬 Key Enzymes & Factors:

📌 Prokaryotic transcription is simpler because RNA polymerase can bind directly to DNA.

🔹 Eukaryotic Transcription Initiation

✔ More complex → RNA polymerase needs transcription factors to help it bind.
✔ Promoter contains a TATA Box (-25 region, TATAAA sequence).
✔ RNA Polymerase II transcribes mRNA.

🔬 Key Enzymes & Factors in Eukaryotic Transcription Initiation:

📌 TFIIH is super important because it both unwinds DNA and starts transcription by phosphorylating RNA Polymerase II!

🔬 4⃣ Step 2: Elongation (Making the RNA)

Goal: RNA polymerase moves along the DNA, adding ribonucleotides (A, U, C, G) to the growing RNA strand.

✔ RNA Polymerase reads the template strand (3' → 5') and synthesizes RNA (5' → 3').
✔ No primer is needed! RNA polymerase can start from scratch.
✔ Uses ribonucleotide triphosphates (rNTPs) as building blocks.
✔ Forms phosphodiester bonds between ribonucleotides.
✔ DNA re-anneals behind RNA polymerase.

🔬 Key Enzymes in Elongation:

📌 Why does RNA Polymerase not need a primer like DNA Polymerase?

• DNA polymerase needs a pre-existing 3' OH group, but RNA polymerase can directly add nucleotides.

🔬 5⃣ Step 3: Termination (Stopping Transcription)

Goal: RNA Polymerase stops and releases the RNA transcript.

🔹 Prokaryotic Termination Mechanisms

✔ Rho-Dependent Termination → Uses the Rho protein (ρ) to detach RNA Polymerase.
✔ Rho-Independent Termination → Forms a GC-rich hairpin loop followed by a U-rich region, which makes RNA polymerase fall off.

🔹 Eukaryotic Termination

✔ RNA Polymerase II continues past the coding region until it reaches the polyadenylation signal (AAUAAA).
✔ The poly-A tail is added, and special enzymes cut the RNA, stopping transcription.

🔬 Key Enzymes & Factors in Termination:

📌 Think of Rho-Independent Termination as RNA "tripping over itself" (hairpin loop) and Eukaryotic Termination as a "cut and run" (AAUAAA signal & CPSF cutting the RNA).

🛠 6⃣ mRNA Processing in Eukaryotes

Before mRNA can leave the nucleus, it undergoes processing to protect it from degradation and help with translation.

✔ 5' Cap Addition → Protects mRNA from degradation and helps ribosome binding.
✔ Splicing → Removes introns (non-coding regions) and joins exons.
✔ Poly-A Tail Addition (~200 Adenines at the 3' End) → Helps with mRNA stability and nuclear export.

🔬 Key Enzymes in mRNA Processing:

📌 Why Do We Need mRNA Processing?
✔ Protects mRNA from exonucleases.
✔ Helps mRNA exit the nucleus.
✔ Ensures proper ribosome recognition for translation.