cell bio 1

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cell bio 1
2013-02-04 17:29:44
cell bio

cell bio
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  1. what does the receptors and signaling mechanisms allow cells to do?
    adapt to environmental conditions
  2. what does the ligand bind?
    receptor to turn it on
  3. what does receptor activate?
    GTP-binding proteins
  4. what happens after enzymes are activated by receptor?
    make second messenger cAMP
  5. what does cAMP activate?
    protein kinases
  6. what do kinases do?
    phosphorylate and active enzymes
  7. what is the job of feedback mechanisms?
    to control molecular composition, growth and differentiation
  8. what are some checkpoints in the cell cycle?
    check for DNA nicks, check for favorable environmental conditions, check for damaged or unduplicated DNA, check for chromosome attachment to mitotic spindle
  9. list the order of the three domains of life in the phylogenetic tree
    archaea, bacteria, and eucarya
  10. explain the prebiotic origin of life
    • occurred about 3.6bya; modern gene sequences are the only surviving evidence
    • simple chemicals, simple RNA's that can store info, complex RNAs with catalytic activity, self replication of catalytic RNAs:
    • -DNA copies of genetic info
    • -encapsulation of nucleic acids in lipid membrane
    • -ribosomes synthesize proteins, which dominate cellular catalysts
  11. what is divergence?
    identical genes diverge from a common ancestor due to a mutation (sister lineages)
  12. Paralogous genes
    diverged as a result of mutations in sister lineages
  13. what are orthologous genes?
    proteins with same functions but found in different species
  14. what are the two scenarios for the origin of eukaryotes?
    • fusion theory: archaea and bacterium are fused and genomes are merged
    • engulfment theory: archaea engulfs the bacterium and the transfer of most bacterial genes to Archaea host
  15. speculation on the origin of organelles in eukaryotes
    prokaryotic extracellular digestive system, fusion of two prokaryotes creates the origin of eukaryote with mitochondria, formation of intracellular digestive system in early eukaryote, formation of membrane organelle and nuclear envelope
  16. what are the functions of membrane?
    compartmentalization, scaffold for reactions, selective permeability barrier, transport, signal transduction, intercelluar interactions, energy transduction. 
  17. how are membrane proteins positioned in the bilayer?
    • membrane proteins have transmembrane domains where the hydrophobic faces of alpha helices and beta sheets associate with membrane lipids
    • beta sheets can form pores with low selectivity
  18. how can we test the mobility of membrane proteins?
    • photobleaching
    • laser trap (immobile proteins do not diffuse freely and particles attached resist displacement
  19. how are membrane proteins solubilized for biochemical studies?
    using nonionic detergents
  20. how to extract membrane proteins?
    integral proteins can be extracted with nondenaturing detergents, peripheral proteins can be extracted using high salt (to disrupt protein-protein interactions
  21. what is the role of peripheral membrane?
    • scaffold for signal transduction
    • skeletal support
    • cell-cell communication and interaction
  22. what is FRAP (Fluorescence recovery after photobleaching) used to measure?
    lateral diffusion rates in membrane
  23. what are the components of reductionist strategy?
    • def of ?
    • inventory of parts
    • concentration (biochm, microscopy)
    • partners (biochm, genetics)
    • rate and equilibrium constants
    • biochemical reconsitition
    • mathematical model
    • physiological tests
  24. what are some cells that you can see without microscopes?
    • Xenopus egg
    • most other cells
    • yeast
    • bacteria
  25. what is the difference between TEM and bright field?
    • You don't have to kill the cell in TEM, but you do in brightfield
    • light microscope is limited by point spread function
  26. what is deconvolution?
    computational method to remove out of focus light from micrographs; take series of laser sections through a cell and use computers to stack images looking at 3-D image of cell, use that go from blurry to high resolution image
  27. describe fluorescence.
    absorption of a photon raises an electron to an excited state; a longer wavelength photon is emitted when the electron falls back to the ground state
  28. fluorescent dye bound to protein
    needs to be microinjected
  29. fluorescent dye bound to lipid need to
    be injected or fused to cell
  30. fluorescent dye bound to nucleic acid
    in situ hybridization in fixed cells
  31. fluorescent dye bound to ligand for cellular molecule
    antibody isolated from animals immunized with target molecule
  32. where does GFP come from (green fluorescent protein)
    • cDNA from jellyfish can be fused to genes for many proteins and expressed in live cells
    • variants are yellow, cyan, red
  33. what does the fluorescent phalloidin bind?
    actin filaments
  34. how to prepare cells from scanning EM?
    fix, dehydrate, coat with metal
  35. how to prepare cells for thin sections on TEM?
    fix, dehydrate, embed, section
  36. how to prepare cells for freeze-fracture and frozen hydrated?
    • freeze rapidly, fracture, metal shadow
    • freeze rapidly, image directly
  37. how to prepare molecules in neg stain, shadowing, frozen hydrated?
    • dry on support film in heavy metal solution
    • dry, evaporate metal film at an angle
    • freeze rapidly, image directly
  38. what makes a good model organism?
    • relevant for the question at hand
    • genome available (not always necessary)
    • ease of maintenance
    • ease of manipulation
    • size
    • ease of analysis (transparency or to find veins)
    • generation time
    • cost and safety
  39. why is E. Coli an excellent model organism?
    • easy to obtain (grows in guts of humans)
    • grows well in lab on borth
    • 20 min duplication time
    • serves to unravel fundamental processes such as: DNA replication, DNA translation into proteins, membrane protein
  40. what is great about brewer's yeast?
    simple eukaryotic, single cell, has mito, small gemoable, amendable to manipulation: genetics, unravel processes such as cell division, basic biochemistry, metabolism, membrane trafficking
  41. why is arabidopsis thaliana a model organism?
    multicellular, has cholorplasts, gorws fast indoors; produces 100s of offspring in weeks, genome is sequenced, understanding food production, crop improvement, ecology
  42. why is nematod worm Caenorhabditis elegans a good model organism?
    • multicellular animal with organs
    • has a gut
    • has neurons
    • develops into 959 cells exactly
    • transparent
    • excellent genetics
    • 70% of genes similar to human genes: study of human diseases
    • understanding programmed cell death, life span analysis, development, neuroscience, behavior and cell division
  43. why is drosophila melanogaster a good model organism?
    • multi-cellular animal
    • has a gut
    • brain/neurons
    • vast number of diff cell types (cell differentiation)
    • larval development (how do legs, wing, etc. end up in right places)
    • genes are similar to human genes (study human diseases)
  44. why s the mouse Mus musculus a model organism?
    • vertebrate animal
    • pigmentless-white mice has transparent skin
    • excellent genetics
  45. what can regenerate the whole body? why?
    planarian flatform, b/c they have stem cells t/o their bodies
  46. what organism can regenerate its limbs?
  47. what are some historic non-genetic model organisms?
    echinoderms; highest of the invertebrates, develops like a vertebrate, biochemistry, live cell imaging, fertilization, cell division, early development, primitive immune system
  48. how is studying model organisms relevant from one species to another?
    b/c of our common descent and the conservation of metabolic and developmental pathways and genetic material over the course of evolution
  49. what is classical genetics?
    make random mutations, identify strains with relevant phenotype, map gene, sequence, gene, identify gene product for functional analysis
  50. what are the molecular inventories?
    • classical genetics
    • genomics
    • isolation of genes and cDNAs
    • biochemical purification of proteins
  51. what is the biochemical purification of proteins?
    develop functional assay, homogenize cells, fractionate until product is pure, obtain sequence, isolate cDNA or gene for functional analysis
  52. how do we isolate genes and cDNAs
    amplify gene by PCR; use DNA probe to clone gene, use expression cloning (functional assay for gene product in bacteria or eukaryotic cell); express gene product in bacteria or eukaryote for functional analysis
  53. how do we do genomics?
    sequence genome or collection of ESTs, identify genes of interest for functional analysis
  54. how do we determine the primary structure of DNA?
    • sequence gene or cDNA, translate protein sequence (DNA sequencing by chain termination with di-deoxynucleotides)
    • sequence protein
  55. how do we determine the subunit composition of DNA?
    • stoichiometry from denaturing gel electrophoresis
    • native molecular weight (ultracentrifuge, gel filtration)
  56. how do we determine the atomic structure of DNA?
    X-ray crystallography, NMR (if <30 kDa)
  57. what are some tests of physiological function?
    • biochemistry means
    • anatomy
    • physiology
  58. what are the biochemistry tests of physiological function?
    reconstitution from purified components
  59. what are the anatomic tests of physiological function?
    localization at the site of action
  60. What are the physiological tests of physiological function?
    • reduced concentration of active proteins (drugs, antibodies, knockout, RNAi, dominant neg mutant)
    • increased concentration of active protein (overexpression)
    • replace native protein with protein having altered properties (classical genetics, homologous recombination
  61. describe the process in coming up with mathematical models
    • postulate the biochemical mechanism
    • write out differential equations describing the pathway
    • stimulate the system behavior using available molecular concentrations, rate and equilibrium constants
    • compare stimulations with data
    • identify defects in model
    • experiment to determine if defects arise from flaws in model or experimental data
    • refine model, predict new properties and test by experimentation
  62. what is the composition of the plasma membrane
    • lipid bilayer (diff types allow for domains in membrane)
    • proteins with function (in, out and through, significant for signaling, interactions, transport, ion, channels, structure, scaffold for carbohydrates)
    • carbohydrates (interactions and signaling)
  63. what are the different types of lipids?
    • phosphoglycerides-phospholipids
    • shingolipids and glycosphingolipids¬†
    • sterolds-cholesterol
  64. what are phosphoglycerides?
    • phophatidyl-choline, ehtanolamine, serine, inositol; differences in different membranes and in domains
    • amphipathic, polar head, nonpolar tail
    • contain choline, phosphate and glycerol with a double bond
    • CDP to CMP
  65. what are the shingolipids?
    • derived from sphingosine, amino alcohol with long hydrocarbon chain and long chain fatty acid attached to sphinogosine
    • glycolipids, gangliosides, sphingomyelin
  66. what is special about the sterols
    • asymmetrically distributed
    • amphipathic b/c of free OH
  67. which direction does GS face in the biological membrane?PS and PI?
    GS face outside; PS and PI face inside
  68. what are rafts rich in?
    cholesterol and sphingolipid
  69. what is the difference between inner membrane and plasma membrane
    • cholestrol makes up about half the lipid in plasma membranes, but is much reduced or absent in intracellular membranes
    • inner mito membrane is protein rich due to high density of respiratory chain enzymes
  70. how are membrane proteins classified?
    transporters, anchors, receptors, enzymes
  71. what are some ways proteins associate with membrane?
    transmembrane (integral protein), monolayer associated, lipid-linked, protein attached
  72. what are the plots that monitor the hydrophobicity of the integral membranes?
    hydropathy plots
  73. what part of the membrane protein is hydrophilic?
    peptide backbone is
  74. what effect underlies the disposition of integral membrane proteins in the bilayer?
    hydrophobic effect
  75. what is the shortest way to span a membrane?
    single alpha hellix about 20 residues @1.5 angstrom per residue
  76. what is the purpose of a membrane skeleton?
    stabilizes the cytoplasmic surface of red blood cell plasma membranes
  77. what causes fragile red blood cells and hemolytic anemia?
    inherited deficiencies of membrane skeleton
  78. channel has low selectivity for molecules with what molecular weight?
    around 600
  79. what kind of side chains do amino acids have?