The Central Dogma
DNA → RNA → Protein — the flow of genetic information
Francis Crick proposed the central dogma of molecular biology in 1958: genetic information flows from DNA to RNA to protein. 'Central' because it is universal to all life, and 'dogma' because it was proposed before complete experimental proof. The dogma has three components: DNA can be replicated (DNA→DNA), DNA can be transcribed (DNA→RNA), and RNA can be translated (RNA→Protein). Reverse transcription (RNA→DNA, performed by retroviruses like HIV) is a recognized exception.
The central dogma explains the fundamental organization of the cell: DNA in the nucleus is the permanent information archive; RNA is the working copy transcribed from specific genes; proteins are the functional molecules that carry out virtually everything the cell does. The separation of archive (nucleus) from manufacturing (cytoplasm, ribosomes) allows exquisite regulation of gene expression — not every gene is copied into every type of cell.
💡 Antibiotics Target Prokaryotic Protein Synthesis
Many antibiotics work by specifically targeting prokaryotic (bacterial) ribosomes (70S) without affecting eukaryotic ribosomes (80S). This selectivity is based on structural differences between bacterial and human ribosomes:
30S subunit inhibitors (block initiation or cause misreading):
• Aminoglycosides (streptomycin, gentamicin, tobramycin): bind 16S rRNA in 30S subunit → cause mRNA misreading → non-functional proteins
• Tetracyclines: block aminoacyl-tRNA binding to the A site of 30S subunit → no elongation
50S subunit inhibitors (block elongation or translocation):
• Macrolides (erythromycin, azithromycin): block translocation → ribosome stalls
• Chloramphenicol: inhibits peptidyl transferase activity → no peptide bond formation
• Linezolid: blocks 70S initiation complex formation
• Clindamycin: blocks translocation (similar to macrolides)
Because mitochondria have 70S ribosomes (endosymbiotic origin from bacteria), some antibiotics — particularly chloramphenicol and aminoglycosides at high doses — can inhibit mitochondrial protein synthesis, causing toxicity to highly metabolic tissues (hearing, kidneys).
Tx
Transcription — copying DNA into RNA
Transcription produces an RNA copy of a gene using the DNA strand as a template. RNA polymerase binds to the promoter sequence (a specific DNA sequence upstream of the gene that signals where transcription should begin), unwinds the DNA, and synthesizes RNA in the 5'→3' direction using the 3'→5' template strand.
Unlike DNA replication, transcription does not require a primer (RNA polymerase can initiate synthesis de novo). Only one strand of DNA (the template strand, also called the antisense or noncoding strand) is used as a template. The other strand (the non-template strand, also called the sense or coding strand) has the same sequence as the mRNA (except U instead of T).
In eukaryotes, transcription occurs in the nucleus. The initial RNA product (pre-mRNA or primary transcript) is processed before export to the cytoplasm. In prokaryotes, transcription and translation occur simultaneously in the same cellular compartment (no nucleus).
Memory trick: Transcription = making mRNA from DNA template. RNA Pol reads the template strand 3'→5' and makes mRNA 5'→3'. The coding strand has same sequence as mRNA (with T replaced by U).
Proc
RNA processing — splicing, capping, polyadenylation
Eukaryotic pre-mRNA undergoes three major processing steps before translation:
5' cap: A 7-methylguanosine cap is added to the 5' end of the pre-mRNA shortly after transcription begins. The 5' cap protects mRNA from degradation by exonucleases and is required for ribosome binding during translation initiation.
3' poly-A tail: After transcription is terminated, the pre-mRNA is cleaved downstream of the stop codon and a string of ~200 adenine nucleotides is added to the 3' end by poly-A polymerase. The poly-A tail protects mRNA from 3' degradation and facilitates export from the nucleus.
Splicing: Most eukaryotic genes contain introns (non-coding intervening sequences) and exons (coding sequences). Introns are removed by the spliceosome (a large RNA-protein complex) and exons are joined together. Alternative splicing — joining exons in different combinations — allows one gene to produce multiple different protein isoforms (the human genome has ~20,000 genes but ~100,000 different proteins, largely due to alternative splicing).
Memory trick: 5' cap = protection helmet. Poly-A tail = protective tail. Splicing = cut out introns (I stay IN the nucleus), keep exons (EXons EXIT). Alternative splicing = same gene, different protein combinations.
Tl
Translation — reading mRNA to make protein
Translation occurs on ribosomes (in eukaryotes, on rough ER for secreted/membrane proteins, or free in the cytoplasm for cytoplasmic proteins). The ribosome reads the mRNA in triplets (codons) from the AUG start codon to a stop codon (UAA, UAG, or UGA).
Transfer RNAs (tRNAs) are adapter molecules that translate codons into amino acids. Each tRNA has a specific anticodon (complementary to a codon) and carries the corresponding amino acid. Aminoacyl-tRNA synthetases attach the correct amino acid to each tRNA.
Three sites on the ribosome: A site (aminoacyl — new tRNA enters), P site (peptidyl — tRNA with growing chain), E site (exit — empty tRNA leaves). Elongation cycle: tRNA with amino acid enters A site → peptide bond formed (peptidyl transferase activity of the large ribosomal subunit) → ribosome translocates 3 nucleotides in 3'→ direction → empty tRNA moves from P to E site and exits → next tRNA enters A site. When a stop codon enters the A site, a release factor binds → the polypeptide is released → the ribosome disassembles.
Memory trick: Ribosomes = A(arrive) → P(peptide) → E(exit). AUG = start (Met). UAA, UAG, UGA = stop. Each codon = 3 nucleotides = 1 amino acid. 64 codons for 20 amino acids = redundancy (degeneracy).
Code
The genetic code — 64 codons, 20 amino acids, 3 stop codons
The genetic code is the relationship between mRNA codons (64 possible triplets of A, U, G, C) and the amino acids they specify. Key features:
Triplet: each codon is 3 nucleotides (because 4³ = 64 > 20 amino acids needed)
Degenerate (redundant): most amino acids are specified by more than one codon (e.g., leucine has 6 codons — UUA, UUG, CUU, CUC, CUA, CUG). This degeneracy is not random — the third position of the codon is most variable (wobble position).
Non-overlapping: each nucleotide is part of only one codon
Commaless: no punctuation between codons — reading frame is maintained from start codon
Universal: (almost) — the same codon specifies the same amino acid in almost all organisms from bacteria to humans. This universality is strong evidence for common ancestry and enables genetic engineering across species.
Memory trick: The genetic code: AUG = Met = START. UAA, UAG, UGA = STOP. 64 codons, 20 amino acids = degeneracy. Universal across all life (almost) = common ancestor.
🔬 Applied Scenario — Mutations in Protein Synthesis and Genetic Disease
Mutations that disrupt protein synthesis cause many of the most important genetic diseases:
A
Sickle cell disease — a missense mutation in hemoglobin. A single nucleotide change (A→T) in codon 6 of the β-globin gene changes the codon from GAG (glutamic acid) to GTG (valine). One amino acid substitution at the protein surface — from a charged, hydrophilic glutamate to a nonpolar, hydrophobic valine — causes hemoglobin to polymerize when deoxygenated → sickle-shaped red blood cells → hemolytic anemia, vaso-occlusive crises, organ damage. This is the paradigmatic example of a missense mutation.
B
Duchenne muscular dystrophy — a frameshift mutation. Most DMD mutations are large deletions in the dystrophin gene that shift the reading frame. A frameshift changes every codon from the mutation point onward → completely different amino acid sequence downstream → premature stop codon → truncated, non-functional protein → absent dystrophin → progressive muscle wasting. Exon skipping therapy (eteplirsen/Exondys 51) uses antisense oligonucleotides to skip the mutated exon → restores reading frame → produces shorter but partially functional dystrophin (like Becker MD).
C
Cystic fibrosis — a deletion mutation causing protein misfolding. The most common CF mutation (ΔF508) is a deletion of 3 nucleotides (in-frame, not a frameshift) encoding phenylalanine at position 508 of the CFTR protein. The resulting protein is one amino acid shorter and misfolds in the ER → targeted for ERAD before reaching the plasma membrane → no functional CFTR chloride channel. Trikafta (elexacaftor/tezacaftor/ivacaftor) corrects ΔF508-CFTR folding and potentiates channel function — directly addressing the molecular consequence of the mutation.
D
Nonsense mutations and stop codon read-through therapy. Nonsense mutations introduce premature stop codons (UAA, UAG, UGA) into the coding sequence → truncated protein → usually non-functional. ~10–15% of inherited disease cases result from nonsense mutations. Aminoglycosides at low concentrations can cause read-through of premature stop codons (misread stop as an amino acid, producing full-length protein). PTC124 (ataluren) is a drug designed to promote read-through of premature stop codons with fewer side effects than aminoglycosides — in clinical trials for DMD and CF caused by nonsense mutations.
📌 Exam Application
Protein synthesis is tested at every level from mechanisms to diseases:
1. Central dogma: DNA → (transcription) → RNA → (translation) → Protein. Reverse transcription (HIV) is the exception.
2. RNA processing (eukaryotes only): 5' cap, 3' poly-A tail, splicing (remove introns, join exons). Alternative splicing → multiple proteins from one gene.
3. Translation sequence: AUG (start/Met) → read codons → UAA/UAG/UGA (stop). A site (arrival) → P site (peptide bond) → E site (exit). Peptidyl transferase = ribosome enzyme.
4. Genetic code features: Triplet, degenerate, non-overlapping, universal (almost). Wobble = third codon position most variable.
5. Antibiotic targets: 30S = aminoglycosides, tetracyclines. 50S = macrolides, chloramphenicol, clindamycin, linezolid.
⚠️ The Most Common Protein Synthesis Mistakes
The template strand is read 3'→5' — but mRNA is synthesized 5'→3'. These are the same process described from two perspectives. RNA polymerase moves along the template strand in the 3'→5' direction, adding nucleotides to the growing 5'→3' RNA chain. Students confuse template strand direction with RNA synthesis direction. The key: the new RNA strand is always synthesized 5'→3', regardless of which DNA strand is the template.
Introns are spliced OUT — exons are kept IN (and exit). The naming is confusing: 'intron' = intervening sequence = stays in the nucleus (removed from mRNA). 'Exon' = expressed sequence = exits the nucleus in the mRNA. Memory trick: 'INtrons stay IN; EXons EXIT (in the mature mRNA).'
The genetic code is read from the mRNA — not from DNA or tRNA. The codon is the mRNA triplet. The anticodon is the complementary tRNA triplet. When given a DNA sequence and asked to find the amino acid sequence, you must: determine which strand is the template → write the complementary mRNA sequence → translate the mRNA codons. Don't try to read the genetic code from DNA directly.
✓ Quick Self-Test
1. What is the central dogma of molecular biology?
2. What three processing steps does pre-mRNA undergo in eukaryotes?
3. What are the three ribosomal sites (A, P, E) and what happens at each?
4. What does 'degenerate genetic code' mean?
5. How do tetracyclines and macrolides each inhibit bacterial protein synthesis?
Answers:
1. The central dogma states that genetic information flows from DNA to RNA to protein: DNA is replicated (DNA→DNA), transcribed (DNA→RNA), and translated (RNA→Protein). Reverse transcription (RNA→DNA, performed by retroviruses) is a recognized exception. The flow is unidirectional — protein sequence information cannot be written back into DNA.
2. Three eukaryotic pre-mRNA processing steps: (1) 5' capping — addition of a 7-methylguanosine cap to protect the mRNA from degradation and enable ribosome binding. (2) 3' polyadenylation — addition of ~200 adenine nucleotides to protect the 3' end and facilitate nuclear export. (3) Splicing — removal of introns and joining of exons by the spliceosome; alternative splicing can produce multiple protein isoforms from one gene.
3. A site (aminoacyl site): incoming aminoacyl-tRNA (carrying the next amino acid) enters and is matched with the current mRNA codon by anticodon-codon base pairing. P site (peptidyl site): tRNA carrying the growing polypeptide chain; peptidyl transferase catalyzes formation of the peptide bond between the growing chain and the new amino acid. E site (exit site): empty tRNA (after transferring its amino acid to the chain) moves here before being released from the ribosome.
4. The genetic code is degenerate (redundant) because most of the 20 amino acids are encoded by more than one codon. With 64 possible codons (4³) and only 20 amino acids plus 3 stop codons, there are 41 'extra' codons — most amino acids have multiple synonymous codons (especially differing at the third/wobble position). This degeneracy buffers against the effects of mutations — a change at the third codon position often specifies the same amino acid (synonymous/silent mutation).
5. Tetracyclines bind the 30S ribosomal subunit and block the A site — preventing aminoacyl-tRNA from entering, so the ribosome cannot receive the next amino acid and elongation stops. Macrolides (erythromycin, azithromycin) bind the 50S ribosomal subunit at the peptide exit tunnel and block translocation — the ribosome cannot move 3 nucleotides along the mRNA after forming each peptide bond, so elongation stalls after the first few amino acids are added.