Exam 3 Flashcards

Translation Overview

  • Translation is the process by which messenger RNA (mRNA) is translated into a polypeptide chain (protein).

  • Occurs in the cytosol where mRNA base pair sequences are translated into amino acid sequences.

  • Nucleotides are read in groups of three known as codons.

  • There are 64 codons made from 4 different bases (A, U, C, G) with each codon specifying either a start, stop signal, or an amino acid.

  • The relationship between a codon and its corresponding amino acid is established by the genetic code.

tRNA Molecule

  • tRNA (transfer RNA) molecules serve as the essential link between codons on mRNA and their corresponding amino acids.

  • Each tRNA is approximately 80 nucleotides in size, featuring an anti-codon region that binds to its complementary codon on mRNA.

Synthesis and Modifications of tRNA

  • tRNA molecules are synthesized by RNA polymerase III (POL III) as large transcripts that are subsequently trimmed at both ends and potentially modified in the middle.

  • These modifications create non-traditional bases (Ψ, Im, Gm, I, T), which contribute to the unique final conformation of the tRNA.

Mechanism of tRNA and Amino Acid Binding

  • Aminoacyl Synthetase is an enzyme that catalyzes the linking of an amino acid to its specific tRNA, forming a high-energy bond.

  • The amino acid-activated tRNA is crucial for peptide bond formation during translation.

  • The reaction consumes energy from ATP, producing AMP and two inorganic phosphates (Pi).

Hydrolytic Editing

  • Prior to catalyzing peptide bond formation, tRNA synthetases can proofread and edit tRNAs, removing incorrect amino acids to maintain fidelity in protein synthesis.

Ribosome Structure and Function

  • The ribosome is a large ribonucleoprotein complex composed of two subunits (large and small) made up of several rRNAs and over 50 proteins.

  • Eukaryotic ribosomes assemble in the nucleolus and are exported into cytosol for protein synthesis. Millions of ribosomes exist per eukaryotic cell.

  • The small subunit contains an mRNA binding site and multiple binding sites for tRNAs, while the large subunit catalyzes peptide bond formation.

Eukaryotic Initiation of Translation

  • The initiation process starts with the initiator aminoacyl-tRNA binding to the small ribosomal subunit.

  • Additional initiation factors (eIFs) bind to guide this small ribosomal subunit to the 5’ cap of mRNA.

  • The small subunit scans the mRNA for the AUG start codon, driving movement powered by ATP hydrolysis.

Leaky Scanning in Eukaryotes

  • When AUG is encountered, initiation factors dissociate, and the large ribosomal subunit binds, starting translation.

  • Sometimes, the first one or two AUGs can be ignored in favor of a later AUG, a phenomenon called leaky scanning, which allows for the synthesis of functionally identical proteins with different localization signals.

Bacterial mRNA and Translation

  • Bacterial mRNAs lack a 5’ cap and instead utilize consensus sequences for small subunit ribosome attachment, which leads to polycistronic mRNAs.

  • This allows for the translation of multiple proteins from a single mRNA strand.

Elongation of the Polypeptide Chain

  • Elongation continues with incoming aminoacyl-tRNAs bringing new amino acids to the A-site of the ribosome.

  • Energy from the breaking of high-energy bonds assists in peptide bond formation, while the large ribosomal subunit catalyzes the reaction, shifting the ribosome down the mRNA template without moving the tRNAs.

Translation Termination

  • Stop codons (UAA, UAG, UGA) signal termination of translation, binding release factors (RFs) that mimic tRNAs.

  • These RFs introduce a water molecule instead of an amino acid, triggering the release of the newly synthesized polypeptide and terminating translation.

Polyribosomes (Polysomes) and Efficiency

  • Polyribosomes are formations of multiple ribosomes translating the same mRNA simultaneously, enhancing the rate of protein synthesis in both prokaryotes and eukaryotes.

  • In eukaryotes, polyribosomes can be formed after transcription is completed, but bacteria begin translation even before transcription ends.

Protein Folding and Quality Control

  • Protein folding begins during synthesis, with the assistance of molecular chaperones (hsp60 and hsp70) that help proteins reach their correct conformations.

  • Proteins that cannot be properly folded are marked for degradation by proteasomes and ubiquitin chains, ensuring cellular quality control by removing defective proteins.

Membrane Structure

Cell Membranes

  • Plasma membrane (in prokaryotes and eukaryotes)

    • Defines cellular space and boundaries.

    • Maintains biochemical and electrical differences.

    • Acts as a conduit for extracellular communication.

  • Organelle membranes (found in eukaryotes only):

    • Define organelle lumens and boundaries.

  • Membrane composition:

    • Lipid bilayer

    • Many inserted and surface-associated proteins

    • A few other types of molecules

Lipid Bilayer

  • Membrane lipids are amphipathic molecules:

    • Contain both polar and non-polar bonds.

  • Membrane lipids have a unique shape:

    • Hydrophilic “head” region (polar) and

    • Hydrophobic “tail” region (non-polar).

  • Membrane lipids spontaneously form bilayers in aqueous solution.

Spontaneous Bilayer Formation

  • Energetically Unfavorable: Lipid micelle (edges exposed to water).

  • Energetically Favorable: Sealed compartment formed by phospholipid bilayer.

Phospholipids

  • Most abundant lipid in cell membranes.

  • Many types of phospholipids:

    • Four major types found in cell membranes.

Four Major Phospholipids

  • Phosphatidyl-ethanolamine

  • Phosphatidyl-serine

  • Phosphatidyl-choline

  • Sphingomyelin

Lipid Bilayer is Fluid

  • Individual lipids diffuse laterally.

  • Rotate rapidly about their axis.

  • Nonpolar tails are extremely flexible.

Cells Regulate Bilayer Fluidity

  • Through:

    • Cis-double bonds in fatty acid tails.

    • Addition of other types of molecules.

  • Allows cells to maintain constant fluidity independent of temperature fluctuations.

Lipid Bilayer Fluidity

  • Affected by the addition of certain non-phospholipid amphipathic molecules, e.g., cholesterol:

    • Interacts with phospholipids to increase rigidity between tails near head region.

    • Lower ends of tails remain flexible and disrupt attractive interactions between phospholipids.

    • This prevents crystallization (freezing).

Lipid Rafts

  • Weak associations between lipids, often include cholesterol and proteins.

    • Lipid rafts drift as a group.

    • Membrane in raft region is often thicker, may serve as a mechanism for holding membrane proteins.

Glycolipids

  • Lipid molecules + attached sugar molecules.

  • Found only on non-cytoplasmic surface (extracellular and intra-organelle).

Glycolipid Functions

  • Protection.

  • Charged glycolipids concentrate ions at cell surface.

  • Cell recognition.

  • Unintended function: may allow bacterial toxins to enter cells.

Membrane Proteins

  • Define much of membrane function.

  • Amount and type of protein in any region of the membrane is highly variable.

  • Membrane proteins often have oligosaccharide chains attached to their non-cytosolic domains (similar to glycolipids).

Mechanisms of Association

  • Transmembrane proteins:

    • Always amphipathic; may be single or multi-pass.

    • Spanning segment often α-helix or β-barrel.

  • Surface proteins:

    • Usually located in cytosolic region; sometimes extracellular.

    • Variety of attachment mechanisms include:

      • α-helix associated with lipid bilayer.

      • Covalent attachment via oligosaccharide chain.

      • Noncovalent attachment to transmembrane protein.

Peripheral vs Integral Proteins

  • Peripheral proteins: Weak noncovalent surface attachment.

  • Integral proteins:

    • Transmembrane or covalently attached to the surface.

    • All peripheral proteins are surface proteins, but not all surface proteins are peripheral.

    • All transmembrane proteins are integral but not all integral proteins are transmembrane.

Examples of Membrane Proteins

  • Spectrin:

    • Cytosolic and noncovalently associated (peripheral) protein found in red blood cells.

    • Associates with cytoskeleton and other proteins to form a flexible mesh, maintaining cell shape (concave red blood cell).

  • Glycophorin:

    • Transmembrane red blood cell protein with a membrane-spanning α helix domain.

    • Extensive extracellular domain composed of oligosaccharide chains, providing adaptability for motility.

  • Bacteriorhodopsin:

    • Multi-pass transmembrane protein functioning as a proton pump in archaea.

    • Contains retinal, responding to photons and enabling mechanical work to pump protons out of the cell.

Membrane Protein Complexes

  • Include several protein subunits such as cytochrome, forming complex cellular structures that participate in various membrane functions.

Glycocalyx

  • Extracellular surface covered with sugars.

  • Covalently attached to:

    • Membrane proteins (glycoproteins)

    • Lipids (glycolipids)

  • Functions:

    • Provides protection against mechanical damage

    • Facilitates cell recognition (as further discussed in Chapter 19).

Proteoglycan

  • A type of glycoprotein in which one or more attached sugars are amino sugars (e.g., glycosaminoglycan).

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