Inhibitors of Transcription — Notes OmpathStudy

Inhibitors of Transcription Transcription inhibitors are compounds that interfere with RNA synthesis by targeting RNA polymerase. Some of these inhibitors are used as antibiotics , while others are natural toxins. 1. Rifamycin (Rifampin) – Inhibitor of Prokaryotic Transcription Produced by : Amycolatopsis rifamycinica (a bacterial species). Targets : Prokaryotic RNA polymerase . Mechanism of Action :Binds to the β-subunit of RNA polymerase . Prevents elongation of the mRNA strand. Inhibits transcription initiation. Clinical Use :Used to treat Mycobacterium tuberculosis (causes tuberculosis ). Effective against other bacterial infections. 2. α-Amanitin – Inhibitor of Eukaryotic Transcription Produced by : Amanita phalloides (death cap mushroom). Targets : Eukaryotic RNA polymerase II . Mechanism of Action : Strongly inhibits RNA polymerase II , which is responsible for synthesizing mRNA . Moderate effect on RNA polymerase III (tRNA synthesis). No effect on RNA polymerase I (rRNA synthesis). Effects :Leads to severe hepatotoxicity (liver damage) . Symptoms include nausea, vomiting, diarrhea , and eventually organ failure . Differences Between Transcription in Prokaryotes and Eukaryotes Post-Transcriptional Modifications in Eukaryotic mRNA 5’ Capping :Addition of a 7-methylguanosine cap at the 5’ end. Protects mRNA from degradation and helps in ribosome binding . Polyadenylation :Addition of a poly-A tail at the 3’ end (about 200 adenine residues). Helps in stability and nuclear export of mRNA. Splicing :Removal of introns (non-coding sequences). Exons (coding regions) are joined together to form mature mRNA . Summary Rifamycin inhibits prokaryotic transcription by binding to RNA polymerase β-subunit . α-Amanitin inhibits eukaryotic transcription by targeting RNA polymerase II . Key differences exist between transcription in prokaryotes and eukaryotes, including location, mRNA processing, regulation, and termination. Eukaryotic mRNA requires post-transcriptional modifications before translation. Translation: Protein Biosynthesis Translation is the second step in gene expression , following transcription. It is the process by which ribosomes synthesize proteins using mRNA as a template. 1. Location of Translation Prokaryotes : Occurs in free ribosomes in the cytoplasm . Eukaryotes : Ribosomes can be free in the cytoplasm or attached to the rough endoplasmic reticulum (RER) , where protein processing occurs . 2. Stages of Translation Protein synthesis occurs in three major steps : (A) Initiation The goal is to bring together the mRNA , tRNA carrying the first amino acid , and the ribosomal subunits . The mRNA contains a sequence of codons (triplets of nucleotides) that dictate which amino acids will be incorporated into the protein. tRNA molecules carry amino acids to the ribosome. The ribosome assembles , with:The small ribosomal subunit binding to the mRNA . The large ribosomal subunit joining later to complete the ribosome. (B) Elongation The ribosome moves along the mRNA , reading codons and adding amino acids in the correct sequence. Each new amino acid is added to the growing protein chain by peptide bond formation . Three key sites in the ribosome: A (Aminoacyl) Site – Binds incoming tRNA carrying an amino acid. P (Peptidyl) Site – Holds tRNA with the growing polypeptide chain . E (Exit) Site – Where empty tRNA exits after donating its amino acid. (C) Termination Occurs when the ribosome reaches a stop codon ( UAA, UAG, UGA ) on the mRNA. A release factor binds to the ribosome, causing it to disassemble , and the newly formed protein is released . 3. Role of tRNA in Translation tRNA (Transfer RNA) molecules bring amino acids from the cytoplasm to the ribosome . tRNA has a unique cloverleaf structure with 73-93 nucleotides . Four distinct regions (lobes) are present: Anticodon Loop – Contains a sequence complementary to the mRNA codon to ensure correct amino acid pairing. Amino Acid Attachment Site – Located at the 3' end , where the correct amino acid binds. 4. Summary of Key Concepts Translation is the process of protein synthesis from mRNA. Takes place in the ribosomes in both prokaryotes and eukaryotes. Involves three stages : Initiation, Elongation, and Termination . tRNA molecules play a crucial role by bringing amino acids to the ribosome and matching codons with their anticodons. Aminoacylation of tRNA (tRNA Charging) 1. Attachment of Amino Acids to tRNA Each tRNA molecule must be attached to its corresponding amino acid to participate in protein synthesis. This attachment process is catalyzed by an enzyme called aminoacyl-tRNA synthetase . There are 20 standard amino acids involved in protein synthesis. Each amino acid has a specific synthetase enzyme that facilitates its attachment to tRNA. 2. Naming of Aminoacyl-tRNA Synthetase The naming of synthetase enzymes is based on the amino acid they attach:Example: The enzyme that attaches leucine to tRNA is called leucyl-tRNA synthetase . Example: The