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BIO 452 Midterm Exam Latest 2025 Cell
Biology Complete Full Length Exam with
Answers California Polytechnic State
University San Luis Obispo, CA
Four types of in vitro cell culture
- Organ Culture: study an organ short term
- Explant Culture: study a piece of tissue w/ different cell types form outgrowth from a single chunk
- Dissociated Cell Culture: cells form a monolayer in a flask (single cell type)
- Organotypic culture: different cell types, recreate an environment realistic to tissue in a regulated environment immunofluorescence microscopy
- allowed for localization of specific proteins within fixed cells
- provided a static image of location
chimeric proteins
- covalently linked protein of interest to fluorescent protein
- visualizing movement of individual proteins on live cells Problem with light microscopy Resolution is too low to examine cells in ultrastructural detail bright field microscopy light shines through specimen epifluorescence microscopy
how to obtain hybridomas
- inject organism with antigen X 2a. some spleen cells make an antibody to antigen 2b. mutant myeloma cells are unable to grow in culture medium, but can survive for long periods of time
- mix and fuse cells (myeloma cells & B-cells)
- transfer to selective medium
- unfused cells die, fused cells survive & grow
- culture surviving cells in separate wells SEM (scanning tunneling microscopy)
- electron beam travels over specimen
- used to view 3D structure, depths
TEM (transmission electron microscopy)
- more flat, 2D representation
- often used in conjunction with antibodies siRNA
- gets rid of mRNA being made ( _____ + mRNA binding)
- w/o mRNA, no protein can be made
- allows for short term decrease in protein expression (or over- expression with a more active promoter) motifs commonly occurring secondary structures
enzyme that de-phosphorylates solvent-accessible surface used to show hydrophobic/hydrophillic interactions what type of forces are involved in secondary structures? hydrogen bonding what type of forces are involved in tertiary structures? non-covalent forces
what three amino acids result in covalent modification of amino acids? tyrosine, serine, & threonine 1st & 2nd Laws of Thermodynamics
- Conservation of energy: energy is neither created nor destroyed
- The entropy of any isolated system always increases enthalpy (H) sum total of all broken and formed bonds How can unfavorable reactions occur? by being coupled to favorable reactions
the hydrolysis of the ATP phosphoanhydride bond is intermediate in bond energy How are membranes able to curve? curvature is due to differences in head group sizes amphipathic both hydrophillic and hydrophobic What factors contribute to a membrane's ability to be maintained in vitro?
- Fatty acid saturation/ unsaturation
- Length of fatty acid tails
- Concentration of cholesterol & cholesterol distribution
Where are nuclear-encoded mRNAs translated on? Cytosolic ribosomes Synthesis of proteins lacking an ER signal sequence is completed on... ...free ribosomes Proteins that contain no targeting sequence are released into the __________ and remain there. cytosol
Four fundamental questions of protein-targeting events
- What is the nature of the signal sequence?
- What is the receptor for the signal sequence?
- What is the structure of the translocon channel that allows the transfer of proteins across the membrane bilayer?
- What is the source of energy that drives unidirectional transfer across the membrane? Ribosomes docked on the rough ER and ribosomes floating in the cytosol are.... the same! What determines whether or not the ribosome binds to the ER? the information in the protein sequence itself
microsomes vesicle fragments that contain bound ribosomes cotranslational translocation simultaneous transport of a secretory protein into the ER as the nascent protein is still bound to the ribosome and being elongated Signal recognition particle (SRP) a cytosolic ribonucleoprotein particle that transiently binds to both the ER signal sequence in a nascent protein as well as the large ribosomal subunit, forming a large complex Signal recognition particle receptor (SRPR)
Where does the protein end up in a lateral exit? Within the lipid bilayer of the membrane (stays in the plasma membrane) exoplasmic face part of membrane that faces the ER lumen cytosolic face part of membrane that faces the cytosol What is both critical to function and maintained after insertion? Orientation
How are membrane proteins classified? By their orientation in the membrane and the types of signals they contain to direct them there topogenic sequences segments within a protein whose sequence, number, & arrangement direct the insertion and orientation of various classes of transmembrane proteins into the ER membrane Type I ER Protein Cytosolic face: C-terminus on a shorter amino acid chain Exoplasmic face: N-terminus on a larger amino acid chain with a signal sequence Example: LDL receptor, insulin receptor
Type I protein insertion
- Ribosome/nascent chain becomes associated with a translocon in the ER membrane
- N-terminal sequence is cleaved
- Chain is elongated until the hydrophobic stop-transfer anchor sequence is synthesized and enters the translocon, preventing the nascent chain from extruding further into the ER lumen stop-transfer anchor sequence (STA sequence) S: stops translating the ribosome T: transfer into lipid bilayer; piece of translocon changes conformation & opens for lateral exit A: anchored by lipids in plasma membrane seen in Type I proteins
Type II protein insertion
- The nascent chain becomes oriented in the translocon with its N-terminal portion toward the cytosol (believed to be mediated by positively charged residues shown N-terminal to the SA sequence)
- As the chain is elongated & extruded to the lumen, the internal SA moves laterally out of the translocon & anchors the chain to the lipid bilayer
- Once protein synthesis is completed, the C-terminus of the polypeptide is released into the lumen, and the ribosomal subunits are released into the cytosol signal-anchor sequence (SA sequence) internal hydrophobic sequence that functions as both an ER signal sequence and a membrane-anchor sequence seen in Type II & Type III proteins
