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Lecture 2: Cell Biology interactive  media  ”video” or ”interactive” 1 Cell biology 2014 (revised 21/1 -14), Note Lecture 2 handout. Alberts et al 5th.

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Presentation on theme: "Lecture 2: Cell Biology interactive  media  ”video” or ”interactive” 1 Cell biology 2014 (revised 21/1 -14), Note Lecture 2 handout. Alberts et al 5th."— Presentation transcript:

1 Lecture 2: Cell Biology interactive  media  ”video” or ”interactive” 1 Cell biology 2014 (revised 21/1 -14), Note Lecture 2 handout. Alberts et al 5th edition Chapter Chapter Chapter A lot of reading! Focus on principles and topics highlighted in the lecture synopsis Ester bond

2 Membranes are primary built from phospholipids Phosphate Glycerol Fatty acid Phosphoglyceride Hydrophilic head Hydrophobic tails The major phospholipid: Variable Fatty acid Lipid bilayer 5 -8 nm thick Biological membranes are lipid bilayers primary composed of amphipathic phospholipids 2 Glycerides (acylglycerols): esters formed from glycerol and fatty acids

3 Packing of amphipathic lipids in water Amphipathic lipids will spontaneously form structures that eliminate the exposure of hydrophobic parts to water - Wedge-shaped lipids form micelles in water - Cylinder-shaped lipids form bilayers, followed by liposome formation 3 H 2 O is a dipole Red: negative Blue: positive

4 Movement of individual lipids within the bilayer Flip-flop (rare) Phospholipids can freely and rapidly (  m/s) diffuse within the monolayer The lipid bilayer is a two-dimensional fluid Similar viscosity as olive oil Spontaneous movements between the two monolayers are rare Rotational and lateral movement (frequent) 4 video 01.2 crawling_amoeba.mov; 13.5 phagocytosis.mov

5 Fatty acid length affects membrane fluidity Long aliphatic carbon chains promote van der Waals interactions  decreased membrane fluidity C=O CH 2 C=O CH 2 C=O CH 2 C=O CH 2 van der Waals Strong interactions Low fluidity Weak interactions High fluidity Long fatty acid tailsShort fatty acid tails 5

6 Fatty acid saturation affects membrane fluidity Phospholipids containing only saturated fatty acids Phospholipids containing a unsaturated fatty acid C=O CH CH 2 C=O CH 2 An unsaturated fatty acid has a kink 6 CH 2 CH Unsaturation's results in steric hindrance  decreased van der Waals interactions  increased membrane fluidity

7 Effect of lipid composition on membrane fluidity - Membrane thickness - Membrane fluidity Shorter fatty acid chains and an increased degree of unsaturation make a thinner and more fluid lipid bilayer 7 - Interactions between fatty acid chains Anim laser_tweezer; Video membrane_fluidity

8 Lipid rafts - clusters of strongly interacting lipids The phospholipid sphingomyelin have long saturated fatty acid tails  strong van der Waals interactions  Formation of a more static lipid environment < 100 nm Lipid rafts are micro-domains of phospholipids with low fluidity 8

9 Inner monolayer (facing the cytosol) Outer monolayer Phosphatidylcholine Phosphatidylethanolamine Phosphatidylserine Sphingomyelin Percentage of membrane lipids Asymmetry of the plasma membrane Phosphatidylinositol, important for cell signaling Lipid raft former Extracellular space molecular_models 10.2-lipids.mov

10 Different types of membrane proteins Integral Peripheral Single-pass  -helix Multi-pass  -helix  -barrel Mono-topic protein Associated to Lipid Integral protein Glycolipid Integral membrane proteins are not tossed into the membrane randomly, but have a specific topology 10

11 Dynamics of membrane proteins Original fluid mosaic model (Singer& Nicolson 1972) Lipid micro-domain (Simons & Ikonen 1997) ~20 % of the plasma membrane Lipid raft 11 Rapid movement of proteins within the lipid bilayer

12 Membrane permeability of different molecules O2O2 CO 2 H2OH2O Ethanol Small uncharged polar molecules Hydrophobic molecules Benzene Na + Charged molecules Ions N C H R C O O H H H + - Amino acids Cl - Large uncharged polar molecules Glucose 12 H+H+

13 Channel proteins Creates a hydrophilic channel through the lipid bilayer that is selective for a particular solute Two types of transmembrane transport proteins Carrier Proteins Binds a “passenger” at one side of membrane and deliver it to the other side From above 13

14 Ion channels Ion OpenClosed Ion AIon B Ion A Most channel proteins are involved in ion transport over the membrane and are therefore called ion channels Ion channels are regulated and ion specific 14

15 Mechanisms behind membrane transport Simple diffusion Facilitated specific diffusion Active transport Energy independent (down-hill) Energy dependent (up-hill) 15 Concentration gradient

16 Different types of active membrane transport Transport of molecules against a concentration gradient requires energy. Cells uses two distinct strategies. ATP-driven pumps Coupled transporters (symporters) “Up-hill” transport of molecule coupled to “down-hill” transport of molecule. The “down-hill” gradient depends on a ATP-driven pump “Up-hill” transport coupled directly to hydrolysis of ATP P ATP ADP + 16

17 Example of active transport - Na + /K + pump Na + K+K+ P ATP ADP Na + P 145 mM 5 mM K+K+ Na + 10 mM 140 mM P K+K+ K+K+ K+K+ K+K+ 1 cycle  10 milliseconds Anim carrier_proteins, Anim Na_K_pump 17

18 Using concentration gradients of Na + and K + K+K+ K+K+ Na + K+K+ K+K+ Active transport of Na + and K + creates concentration gradients The Na + gradient provides the energy for “up-hill transport” Glucose Glucose Coupled transport of sucrose into the cytosol The ATP driving the Na + /K + pump is the energy source for concentrating sugars and amino acids within cells 18

19 Example of trans-cellular transport by a symporter Glucose Na + Glucose Active transport: Na + driven glucose symport (“cotransporter”) 3. Na + Passive transport: facilitated “specific” diffusion of glucose to blood 3. Na + /K + pump establish Na + gradient Na + Glucose Intestinal lumen Glucose Blood vessels K+K+ K+K+ K+K+ K+K+ K+K+ ATP Anim glucose_uptake 19

20 CompartmentMain function Cytosol Protein synthesis, metabolism Nucleus DNA & RNA synthesis Endoplasmic Lipid synthesis, synthesis of proteins that reticulum (ER) enters the secretory pathway Golgi Sorting and packaging for delivery to cell Lysosome Protein degradation Mitochondrion ATP production surface or lysosome 20 Compartments/organelles of eukaryotic cells

21 The nucleus – the instruction book of the cell Nuclear pore rRNA + proteins DNA replication Transcription  mRNA, rRNA and tRNA Ribosome subunit assembly 3-10  m Nuclear processes: 21

22 One reason for a nucleus in eukaryotes Transcription Translation mRNA processing Transcription Translation ProkaryoteEukaryote In eukaryotes mRNA has to be processed prior to initiation of translation, which requires spatial separation of transcription and translation ( Note cloning of an ORF  cDNA synthesis ) 22

23 Transport in and out of the nucleus Nuclear pore Nuclear pore rRNA mRNA tRNA Protein synthesis in the cytosol DNA replication Transcription 2. 23

24 The nuclear pore complex (NPC) Inner nuclear membrane Outer nuclear membrane 120 nm Annular subunit; the gatekeeper Proteins less than 60 kDa can diffuse ”freely” between cytosol and nucleus A typical cell contains nuclear pore complexes 24

25 Nuclear import of proteins (>60kD) NLS NC NC NC Nuclear Localization Sequence (NLS) = sequence in a protein that mediates nuclear uptake Could be localized anywhere in the protein N NC LS NLS NC Even distant apart in the primary structure of the protein Which becomes adjacent in the folded protein 25

26 The process of facilitated nuclear protein import Nuclear import receptor (importin) NLS NLS 4. Association of target protein and nuclear import receptor in the cytosol Binding to the nuclear pore complex mediated by the nuclear import receptor ”Walking” through the gate-keepers of the pore Dissociation of target protein and nuclear import receptor inside the nucleus 26

27 The nuclear import cycle CytosolNucleus Importin NLS Importin NLS Importin GTP Ran Importin GTP Ran GTP Ran NLS Importin Ran GDP Importin NLS Ran GDP +Pi <60 kDa 27

28 The driving forces behind nuclear import CytosolNucleus Importin NLS Importin GTP Ran GTP Ran NLS Importin NLS Importin NLS Importin GTP Ran GTP GDP Energy cost! Ran GDP Ran GDP <60 kDa 28 Video 02.3-brownian_motion.mov

29 G protein Directionality in nuclear import – the Ran cycle Cytosol Nucleus GTP Ran GTP Ran GDP Ran GDP Ran-GAP Ran-GEF GTP G protein GDP GTPase Activating Protein (GAP) Guanine-nucleotide Exchange Factor (GEF) Pi << GDPGTP 29

30 Nuclear export NLSNES Nuclear export of proteins is mediated by an intrinsic Nuclear Export Signal (NES). Proteins with NES include: Small protein that should not be nuclear Protein that shuttle between cytosol and nucleus Export of mRNA is dependent on successful splicing NSEProteins responsible for splicing NSE Spliced mRNA ready for nuclear export Splicing; removal of introns from mRNA 30 Video 12.2-nuclear_import.mov


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