π Transcription & Genetic Code
Central Dogma of Molecular Biology
Proposed by Francis Crick (1958). The flow of genetic information follows a specific direction:
DNA β (Transcription) β RNA β (Translation) β Protein
Exceptions to the central dogma:
- Reverse transcription: RNA β DNA (by reverse transcriptase in retroviruses, e.g., HIV). Discovered by Howard Temin and David Baltimore β Nobel Prize 1975.
- RNA replication: RNA β RNA (RNA viruses, e.g., influenza, SARS-CoV-2 β by RNA-dependent RNA polymerase).
- Prions: Protein β Protein (infectious proteins, no nucleic acid; cause mad cow disease/BSE).
Types of RNA
| Type | Full Name | % of Total RNA | Function |
|---|---|---|---|
| rRNA | Ribosomal RNA | ~80% | Structural component of ribosomes; catalytic (ribozyme) in peptide bond formation |
| tRNA | Transfer RNA | ~15% | Carries amino acids to ribosome; adaptor molecule |
| mRNA | Messenger RNA | ~3-5% | Carries genetic code from DNA to ribosome; template for protein synthesis |
TIP
Despite being most discussed, mRNA is the least abundant (~3-5%). rRNA dominates at 80% because ribosomes are needed in enormous quantities.
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Central Dogma of Molecular Biology
Proposed by Francis Crick (1958). The flow of genetic information follows a specific direction:
DNA β (Transcription) β RNA β (Translation) β Protein
Exceptions to the central dogma:
- Reverse transcription: RNA β DNA (by reverse transcriptase in retroviruses, e.g., HIV). Discovered by Howard Temin and David Baltimore β Nobel Prize 1975.
- RNA replication: RNA β RNA (RNA viruses, e.g., influenza, SARS-CoV-2 β by RNA-dependent RNA polymerase).
- Prions: Protein β Protein (infectious proteins, no nucleic acid; cause mad cow disease/BSE).
Types of RNA
| Type | Full Name | % of Total RNA | Function |
|---|---|---|---|
| rRNA | Ribosomal RNA | ~80% | Structural component of ribosomes; catalytic (ribozyme) in peptide bond formation |
| tRNA | Transfer RNA | ~15% | Carries amino acids to ribosome; adaptor molecule |
| mRNA | Messenger RNA | ~3-5% | Carries genetic code from DNA to ribosome; template for protein synthesis |
TIP
Despite being most discussed, mRNA is the least abundant (~3-5%). rRNA dominates at 80% because ribosomes are needed in enormous quantities.
tRNA Structure (Cloverleaf Model)
- Described by Robert Holley (1965) β first nucleic acid to be fully sequenced (yeast alanine-tRNA). Nobel Prize 1968.
- L-shaped 3D structure; Cloverleaf shape in 2D.
- ~75-90 nucleotides long.
Key features:
- Acceptor arm (CCA arm) β 3' end sequence CCA; amino acid binds here. Universal across all tRNAs.
- Anticodon arm β contains the anticodon (3 bases complementary to mRNA codon).
- D arm (DHU arm) β contains dihydrouridine; recognition site for aminoacyl-tRNA synthetase.
- TΟC arm β contains ribothymidine and pseudouridine (Ο); helps ribosome binding.
- Variable arm β varies in size; used for tRNA classification.
NOTE
tRNA has several unusual/modified bases: pseudouridine (Ο), dihydrouridine (D), inosine (I). These are important for tRNA stability and function.
Image Generation Prompt
A cloverleaf diagram of tRNA showing: acceptor stem with CCA-3' end where amino acid attaches, anticodon loop with three anticodon bases, D arm (DHU loop), TΟC arm, and variable arm. Label all four arms/loops and 5' and 3' ends. Show amino acid attachment site. Scientific illustration.
Transcription (DNA β RNA)
Transcription copies genetic information from one strand of DNA into an RNA molecule.
- Catalyzed by RNA polymerase.
- Occurs in the 5'β3' direction.
- No primer needed (unlike DNA replication).
Key Terms
| Term | Definition |
|---|---|
| Template strand (Antisense) | DNA strand read by RNA polymerase (3'β5'); complementary to mRNA |
| Coding strand (Sense/Non-template) | Same sequence as mRNA (with T instead of U); NOT read by RNA polymerase |
| Promoter | DNA sequence where RNA polymerase binds to initiate transcription |
| Terminator | DNA sequence signaling end of transcription |
| Transcription unit | Promoter + Structural gene + Terminator |
Transcription in Prokaryotes
RNA Polymerase in E. coli
- Single type of RNA polymerase synthesizes all RNA types (mRNA, tRNA, rRNA).
- Holoenzyme = Core enzyme + Sigma (Ο) factor.
- Core enzyme: Ξ±βΞ²Ξ²'Ο
- Ο factor: Required for promoter recognition; most common = Οβ·β°.
- After initiation, sigma factor dissociates β core enzyme continues elongation alone.
Steps of Prokaryotic Transcription
1. Initiation:
- RNA polymerase holoenzyme binds to the promoter.
- Pribnow box (-10 region): consensus TATAAT; DNA unwinds here (A-T pairs = 2 H-bonds, easier to separate).
- -35 region: consensus TTGACA; first contact point of RNA polymerase.
- +1 site: Transcription start; first nucleotide usually a purine (A or G).
- DNA unwinds locally (~17 bp) β transcription bubble / open complex.
2. Elongation:
- Sigma factor released after ~8-10 nucleotides.
- Core enzyme moves along template strand 3'β5'.
- RNA synthesized 5'β3' (NTPs: ATP, UTP, GTP, CTP).
- Speed: ~40-50 nucleotides/second in prokaryotes.
3. Termination:
Two termination mechanisms
**Rho-independent (Intrinsic):** Terminator sequence forms a **GC-rich hairpin (stem-loop)** followed by a run of **U's** in the mRNA. Hairpin causes polymerase to pause; weak rU-dA bonds break β RNA releases.Rho-dependent: Rho protein (Ο) β a helicase that moves along mRNA 5'β3'. When RNA polymerase pauses at the terminator, rho catches up and unwinds the RNA-DNA hybrid β RNA released.
Transcription in Eukaryotes
Three RNA Polymerases
| RNA Polymerase | Location | Product | Sensitivity to Ξ±-amanitin |
|---|---|---|---|
| RNA Pol I | Nucleolus | rRNA (28S, 18S, 5.8S) | Insensitive |
| RNA Pol II | Nucleoplasm | mRNA (and snRNA) | Most sensitive |
| RNA Pol III | Nucleoplasm | tRNA, 5S rRNA, snRNA | Moderately sensitive |
WARNING
Ξ±-amanitin β toxin from Amanita phalloides (death cap mushroom); inhibits RNA Pol II most strongly. Stops mRNA production β protein synthesis halts β organ failure.
Eukaryotic Promoter Elements
- TATA box (Hogness box) β TATAAA at -25 to -30. Analogous to prokaryotic Pribnow box.
- CAAT box β at -70 to -80; determines frequency of initiation.
- GC box β at -110; associated with housekeeping genes.
- Enhancers β can be thousands of bp upstream or downstream; increase transcription rate.
- Transcription factors (TFs) β proteins helping RNA Pol II bind promoter (e.g., TFIID binds TATA box via TBP).
Post-Transcriptional Modifications (RNA Processing)
In eukaryotes, the primary transcript (hnRNA β heterogeneous nuclear RNA / pre-mRNA) undergoes modifications before becoming mature mRNA. Prokaryotic mRNA has NO such modifications β it is translated even while being transcribed (coupled transcription-translation).
1. 5' Capping
- Addition of 7-methylguanosine (mβ·G) cap to the 5' end via a 5'-5' triphosphate linkage.
- Functions: Protects mRNA from degradation; ribosome recognition; aids nuclear export.
2. 3' Polyadenylation (Poly-A Tail)
- Addition of 100-250 adenine nucleotides (poly-A tail) by poly-A polymerase.
- Signal: AAUAAA sequence (polyadenylation signal).
- Functions: Protects from degradation; mRNA export; translation efficiency; stability.
3. RNA Splicing
- Removal of introns (non-coding/intervening sequences) and joining of exons (expressed/coding sequences).
- Carried out by the spliceosome β complex of snRNPs and proteins.
- snRNAs: U1, U2, U4, U5, U6 β recognize splice sites.
- Lariat structure β branched intermediate formed during splicing.
- Splice site sequences: 5' splice site = GU; 3' splice site = AG.
- GT-AG rule (DNA level: GU-AG at RNA level) β introns begin with GU and end with AG.
Split Genes (Interrupted Genes)
- Discovered by Richard Roberts and Phillip Sharp (1977) β Nobel Prize 1993.
- Eukaryotic genes have exons interspersed with introns.
- Prokaryotic genes are generally continuous (no introns).
Alternative Splicing
- One gene β multiple mRNAs (and thus multiple proteins) by including/excluding different exons.
- Example: Drosophila Dscam gene β over 38,000 different mRNAs.
- Explains how ~20,000 human genes encode >100,000 different proteins.
Genetic Code
Properties
The genetic code was deciphered by Marshall Nirenberg, Har Gobind Khorana, and Robert Holley β Nobel Prize 1968.
| Property | Description |
|---|---|
| Triplet | Each codon = 3 consecutive nucleotides β one amino acid |
| Non-overlapping | Codons read sequentially without sharing nucleotides |
| Commaless (Continuous) | No gaps or punctuation between codons |
| Non-ambiguous | Each codon codes for only one amino acid |
| Degenerate (Redundant) | Multiple codons can code for the same amino acid (e.g., 6 codons for Leu/Ser/Arg) |
| Universal | Same code in almost all organisms (few exceptions in mitochondria) |
| Unidirectional | Read in 5'β3' direction on mRNA |
NOTE
Non-ambiguous β Degenerate. Non-ambiguous = one codon β one amino acid. Degenerate = one amino acid β multiple codons. These coexist.
Special Codons
| Codon | Function |
|---|---|
| AUG | Start codon (codes for Methionine; in prokaryotes = N-formyl methionine fMet) |
| UAA | Stop codon β "Ochre" |
| UAG | Stop codon β "Amber" |
| UGA | Stop codon β "Opal/Umber" |
- 64 codons total (4Β³ = 64).
- 61 sense codons for 20 amino acids.
- 3 stop (nonsense) codons.
- Methionine (AUG) and Tryptophan (UGG) β coded by only one codon each (non-degenerate).
Wobble Hypothesis
- Proposed by Francis Crick (1966).
- The 3rd position of the codon and 1st position of the anticodon can form non-standard (wobble) base pairs.
- Allows a single tRNA to recognize more than one codon (explains degeneracy with fewer than 61 tRNAs).
- Wobble pairs: G-U, I-U, I-C, I-A (I = inosine).
Key Points to Remember
- Central dogma: DNA β RNA β Protein (Crick, 1958); exceptions: reverse transcription (HIV), RNA replication (RNA viruses), prions
- tRNA: cloverleaf model (Holley, 1965; Nobel 1968); 3' end = CCA; anticodon loop recognizes codon
- Prokaryotes: one RNA polymerase; Οβ·β° for promoter recognition; Pribnow box (TATAAT, -10), -35 region (TTGACA)
- Eukaryotes: 3 RNA polymerases; RNA Pol II β mRNA; TATA box (-25 to -30); Ξ±-amanitin inhibits RNA Pol II most
- RNA processing: 5' mβ·G cap, poly-A tail (100-250 A's, signal AAUAAA), splicing (spliceosome, GT-AG rule)
- Split genes: Roberts & Sharp (1977), Nobel 1993; exons = expressed, introns = intervening
- Genetic code: 64 codons, 61 sense + 3 stop (UAA/UAG/UGA); AUG = start; degeneracy; universal; wobble (Crick, 1966)
- Met (AUG) and Trp (UGG) = only amino acids with single codon
Summary Cheat Sheet
| Concept / Topic | Key Details / Explanation |
|---|---|
| Central Dogma | DNA β RNA β Protein; proposed by Francis Crick (1958) |
| Exceptions to central dogma | Reverse transcription (RNA β DNA; retroviruses/HIV; Temin & Baltimore, Nobel 1975) RNA replication (RNA β RNA; RNA viruses) Prions (Protein β Protein; no nucleic acid) |
| rRNA | ~80% of total RNA; structural/catalytic component of ribosomes |
| tRNA | ~15%; carries amino acids; adaptor molecule |
| mRNA | ~3-5% (least abundant); template for protein synthesis |
| tRNA structure | Cloverleaf (2D) / L-shaped (3D); described by Robert Holley (1965), Nobel 1968; ~75-90 nucleotides |
| tRNA key features | CCA at 3' end (amino acid attachment) Anticodon arm (recognizes mRNA codon) D arm (dihydrouridine) TΟC arm (ribosome binding) |
| Template strand | Antisense; read by RNA polymerase 3'β5' |
| Coding strand | Sense; same sequence as mRNA (T instead of U); NOT read by polymerase |
| Transcription unit | Promoter + Structural gene + Terminator |
| Prokaryotic RNA polymerase | Single type; holoenzyme = core (Ξ±βΞ²Ξ²'Ο) + Ο factor (Οβ·β° most common) |
| Pribnow box | TATAAT at -10 region; DNA unwinds here (A-T rich) |
| -35 region | TTGACA; first contact point of RNA polymerase |
| Prokaryotic termination | Rho-independent (GC hairpin + poly-U) or Rho-dependent (Ο helicase) |
| Eukaryotic RNA Pol I | Nucleolus; makes rRNA (28S, 18S, 5.8S); insensitive to Ξ±-amanitin |
| Eukaryotic RNA Pol II | Nucleoplasm; makes mRNA; most sensitive to Ξ±-amanitin |
| Eukaryotic RNA Pol III | Nucleoplasm; makes tRNA, 5S rRNA; moderately sensitive |
| Ξ±-amanitin | Toxin from Amanita phalloides; inhibits RNA Pol II most strongly |
| TATA box (Hogness box) | TATAAA at -25 to -30; eukaryotic promoter element |
| 5' Capping | 7-methylguanosine (mβ·G) cap; protects mRNA; ribosome recognition |
| 3' Polyadenylation | 100-250 adenines (poly-A tail); signal = AAUAAA |
| RNA Splicing | Removal of introns, joining of exons; by spliceosome (snRNPs) |
| GT-AG rule | Introns begin with GU and end with AG (at RNA level) |
| Split genes discovered by | Richard Roberts & Phillip Sharp (1977); Nobel 1993 |
| Alternative splicing | One gene β multiple mRNAs/proteins; Drosophila Dscam β >38,000 mRNAs |
| Genetic code deciphered by | Nirenberg, Khorana, Holley; Nobel 1968 |
| Total codons | 64 (4Β³); 61 sense + 3 stop |
| Start codon | AUG (Methionine; fMet in prokaryotes) |
| Stop codons | UAA (Ochre), UAG (Amber), UGA (Opal) |
| Single-codon amino acids | Met (AUG) and Trp (UGG) only |
| Genetic code properties | Triplet, non-overlapping, commaless, non-ambiguous, degenerate, universal, unidirectional (5'β3') |
| Wobble hypothesis | Crick (1966); 3rd codon position allows non-standard pairing; fewer tRNAs needed |
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