Central Dogma of Molecular Biology

Overview of Genetic Information Flow

The central dogma of molecular biology describes the directional flow of genetic information within a cell: DNA is replicated, transcribed into RNA, and then translated into protein. This process is fundamental to all living organisms and underpins gene expression and inheritance.

• Replication: DNA is duplicated by DNA polymerase to ensure genetic continuity during cell division.

• Transcription: Genetic information is transferred from DNA to RNA by RNA polymerase.

• Translation: Ribosomes use mRNA as a template to synthesize polypeptides (proteins).

• Exceptions: Retroviruses can reverse transcribe RNA into DNA.

Genetic Material and Expression

• Genetic material: DNA is responsible for inheritance and directs protein synthesis.

• Gene expression: The process by which genetic information is used to synthesize proteins, determining phenotype.

Transcription – RNA Synthesis

Transcriptional Units and Regulation

Transcription is the first step in gene expression, where RNA is synthesized from a DNA template. Genes may be constitutive (always expressed) or regulated (expressed under specific conditions).

• Transcriptional units: Segments of DNA transcribed into a single RNA molecule.

• Monocistronic mRNA: Encodes a single gene.

• Polycistronic mRNA: Encodes multiple genes (operons), common in prokaryotes.

Transcription in Prokaryotes vs Eukaryotes

• Prokaryotes: Replication, transcription, and translation occur in the cytoplasm; transcription and translation are coupled; polycistronic mRNA is common.

• Eukaryotes: Replication and transcription occur in the nucleus; mRNA is processed (5’ cap, splicing, 3’ poly-A tail) and exported to the cytoplasm for translation; monocistronic mRNA is typical.

Transcription Mechanism

• Initiation: Sigma factor recognizes promoter, recruits RNA polymerase, forms transcription bubble, and initiates RNA synthesis.

• Elongation: RNA polymerase adds ribonucleotides to the 3’ end of the growing RNA strand.

• Termination: Can be Rho-dependent (Rho protein releases RNA polymerase) or intrinsic (hairpin loop formation in RNA).

RNA Polymerase and Sigma Factors

• RNA polymerase: Core enzyme synthesizes RNA; sigma factor recognizes promoter regions.

• Promoter: Contains conserved sequences (Pribnow box at -10, -35 element) recognized by sigma factors.

RNA Products and Types

• mRNA: Blueprint for protein synthesis (codons).

• tRNA: Carries amino acids for translation (anticodon).

• rRNA: Catalytic RNA component of ribosomes.

Comparison of Transcription in Bacteria and Eukaryotes

• Bacteria: Monocistronic or polycistronic mRNA, no 5’ cap or poly-A tail, no splicing.

• Eukaryotes: Exclusively monocistronic mRNA, 5’ cap, poly-A tail, splicing of pre-mRNA.

The Genetic Code

Triplet Code and Codon Dictionary

The genetic code is a set of rules by which information encoded in mRNA is translated into proteins. It is universal, unambiguous, and degenerate.

• Codon: Three-nucleotide sequence in mRNA specifying an amino acid.

• Degeneracy: Multiple codons can specify the same amino acid.

• Start codon: AUG (methionine or formyl methionine in prokaryotes).

• Stop codons: UAA, UAG, UGA.

• Wobble effect: Third base in codon is less important for tRNA binding.

Reading Frames and Mutations

• Reading frame: Uninterrupted sequence of nucleotides translated into a polypeptide.

• Frame shift mutations: Addition or deletion of nucleotides alters the reading frame, affecting protein synthesis.

Translation – Protein Synthesis

Translation Mechanism and Machinery

Translation is the process by which ribosomes synthesize proteins using mRNA as a template. It involves tRNA, aminoacyl-tRNA synthetase, and ribosomes.

• tRNA: Carries amino acids to ribosome; has acceptor stem and anticodon.

• Aminoacyl-tRNA synthetase: Enzyme that attaches correct amino acid to tRNA.

• Ribosome: Composed of large and small subunits; catalyzes peptide bond formation.

• Binding sites: A (aminoacyl), P (peptidyl), E (exit).

Translation Steps

• Initiation: Small ribosomal subunit binds to mRNA (Shine-Dalgarno sequence in prokaryotes), initiator tRNA binds to P site, large subunit joins.

• Elongation: Cycles of aminoacyl-tRNA binding, peptide bond formation, and translocation; GTP-driven.

• Termination: Stop codon recognized by release factors, polypeptide released, translation machinery dissociates.

Translation in Prokaryotes vs Eukaryotes

• Prokaryotes: Monocistronic and polycistronic mRNA, translation starts at multiple sites.

• Eukaryotes: Monocistronic mRNA, translation starts at a single site.

Protein Maturation, Folding, and Sorting

Post-Translational Modifications

Proteins undergo modifications after translation to achieve functional maturity, stability, and proper localization.

• Deformylation: Removal of N-formyl group.

• Methionine excision: Removal of N-terminal methionine.

• Covalent additions: Phosphorylation, glycosylation, methylation, lipidation, acetylation, etc.

Protein Folding – Chaperone Proteins

• Chaperones: Assist in proper folding of proteins; ATP-dependent.

• Types: Trigger factors, DnaK/DnaJ, GroEL/GroES, heat-shock and cold-shock proteins.

Protein Sorting and Export

• Sorting: Proteins are directed to cytosol, membrane, periplasm, outer membrane, or extracellular environment.

• Signal sequence: N-terminal peptide directs protein to export pathways.

• TAT signal peptide: Contains two arginine residues (RR).

Translocases and Secretion Systems

Sec System

• SRP-SecYEG: Signal recognition particle targets ribosome to SecYEG translocon for co-translational translocation.

• SecA-SecYEG: SecA targets unfolded protein to SecYEG for post-translational translocation; energy from ATP and PMF.

TAT System (Twin Arginine Translocase)

• Transports folded proteins with TAT signal sequence.

• Energy: Proton-motive force (PMF).

Secretion Systems in Gram-Negative Bacteria

• Two-step systems: Type V (autotransporters), Type II (secretion pore).

• One-step systems: Type I (ABC transporter), Type III (injectisome), Type IV (DNA transfer), Type VI (needle-like projection).

Additional Info: Meselson & Stahl Experiment

DNA Replication Mechanism

The Meselson & Stahl experiment used heavy and light nitrogen isotopes to demonstrate the semi-conservative mechanism of DNA replication.

• Semi-conservative replication: Each new DNA molecule contains one old and one new strand.