DNA Replication: Molecular Biology

• Molecular biology explains life processes at the molecular level, focusing on the structure, function, and interactions of macromolecules (proteins and nucleic acids) within cells

DNA Replication

• DNA replication is the process of producing two identical replicas from one original DNA molecule
• It ensures genetic information is accurately passed from one generation to the next
• The process is described as semi-conservative because each new DNA molecule consists of one original (template) strand and one newly synthesized strand
• Replication starts at specific sites called origins of replication, where the DNA double helix unwinds
• Helicase unwinds the DNA double helix, creating a replication fork
• Single-strand binding proteins (SSBPs) prevent the separated strands from re-annealing
• DNA polymerase synthesizes new DNA strands by adding nucleotides to the 3' end of a pre-existing strand
• DNA polymerase requires a primer to start synthesis, which is synthesized by primase
• The leading strand is synthesized continuously in the 5' to 3' direction towards the replication fork
• The lagging strand is synthesized discontinuously in short fragments (Okazaki fragments) in the 5' to 3' direction away from the replication fork
• Okazaki fragments are joined together by DNA ligase
• DNA polymerase also has proofreading capabilities, correcting errors during replication
• Topoisomerases relieve strain ahead of the replication fork by breaking, swiveling, and rejoining DNA strands

Transcription

• Transcription is the process of synthesizing RNA from a DNA template
• It is the first step in gene expression
• RNA polymerase binds to a specific DNA sequence called the promoter to initiate transcription
• In eukaryotes, transcription factors help RNA polymerase bind to the promoter
• RNA polymerase unwinds the DNA and synthesizes an RNA molecule complementary to the template strand (antisense strand) in the 5' to 3' direction
• The RNA molecule is synthesized using the coding strand (sense strand) as a reference, with uracil (U) replacing thymine (T)
• Transcription continues until RNA polymerase reaches a terminator sequence
• In eukaryotes, the primary RNA transcript (pre-mRNA) undergoes processing
• RNA splicing: introns (non-coding regions) are removed, and exons (coding regions) are joined together
• 5' capping: a modified guanine nucleotide is added to the 5' end
• 3' polyadenylation: a poly(A) tail (a string of adenine nucleotides) is added to the 3' end
• These modifications protect the mRNA from degradation and enhance translation

Translation

• Translation is the process of synthesizing a polypeptide chain (protein) from mRNA
• It occurs on ribosomes in the cytoplasm
• mRNA binds to the ribosome, and translation begins at the start codon (AUG)
• Transfer RNA (tRNA) molecules bring specific amino acids to the ribosome
• Each tRNA has an anticodon that is complementary to a specific mRNA codon
• The ribosome moves along the mRNA, codon by codon
• As each codon is read, the corresponding tRNA brings the appropriate amino acid to the ribosome
• Amino acids are joined together by peptide bonds, forming a polypeptide chain
• Translation continues until the ribosome reaches a stop codon (UAA, UAG, or UGA)
• Upon reaching a stop codon, a release factor binds to the ribosome, and the polypeptide chain is released
• The polypeptide chain then folds into its functional three-dimensional structure

Genetic Engineering

• Genetic engineering involves the modification of an organism's genetic material using biotechnology
• Restriction enzymes cut DNA at specific sequences, creating fragments with sticky or blunt ends
• DNA ligase joins DNA fragments together
• Plasmids are small, circular DNA molecules found in bacteria, often used as vectors to carry foreign DNA into cells
• Recombinant DNA is DNA that has been artificially combined from different sources
• Transformation is the process of introducing foreign DNA (e.g., a recombinant plasmid) into a host cell
• Polymerase Chain Reaction (PCR) is a technique used to amplify specific DNA sequences
• Gel electrophoresis separates DNA fragments based on size
• DNA sequencing determines the nucleotide sequence of a DNA molecule
• Genetic engineering applications include:
• Production of recombinant proteins (e.g., insulin)
• Gene therapy
• Genetically modified organisms (GMOs)
• Disease diagnostics

Cell Signaling

• Cell signaling refers to the process by which cells communicate with each other to coordinate their activities
• Signaling involves:
• Signal reception: a signaling molecule (ligand) binds to a receptor protein on the cell surface or inside the cell
• Signal transduction: the signal is converted into a form that can bring about a cellular response, often involving a cascade of protein modifications (phosphorylation)
• Cellular response: a change in the cell's activity, such as gene expression, metabolism, or cell shape
• Types of cell signaling:
• Endocrine signaling: hormones are secreted into the bloodstream and travel to target cells throughout the body
• Paracrine signaling: signaling molecules act on nearby cells
• Autocrine signaling: a cell signals itself
• Direct contact: cells communicate through direct physical contact
• Receptor types include:
• G protein-coupled receptors (GPCRs)
• Receptor tyrosine kinases (RTKs)
• Ligand-gated ion channels
• Secondary messengers (e.g., cAMP, calcium ions) amplify the signal within the cell

Protein Synthesis

• Protein synthesis is the process of creating protein molecules
• It includes transcription and translation
• Transcription:
• DNA is transcribed into mRNA in the nucleus
• RNA polymerase binds to the promoter region of a gene
• mRNA is processed (splicing, capping, polyadenylation) before leaving the nucleus
• Translation:
• mRNA binds to ribosomes in the cytoplasm
• tRNA molecules bring amino acids to the ribosome
• Codons on mRNA are matched with anticodons on tRNA
• Amino acids are linked by peptide bonds to form a polypeptide chain
• Post-translational modifications:
• Polypeptides may undergo folding, glycosylation, phosphorylation, or other modifications
• Proteins are transported to their final destination in the cell
• Regulation of protein synthesis:
• Gene expression can be regulated at various steps (transcription, translation, post-translation)
• Regulatory proteins (transcription factors) can enhance or inhibit transcription
• RNA interference (RNAi) can block translation or degrade mRNA