EXAM 1 STUDY GUIDE
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This 21 page Study Guide was uploaded by Luke Holden on Sunday September 11, 2016. The Study Guide belongs to BIOL 4610 at Clemson University taught by Susan Chapman in Fall 2016. Since its upload, it has received 107 views.
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Date Created: 09/11/16
Membrane lipids LIPID STRUCTURE BASE IDENTIFIERS Phospholipid Polar head group Glycerol (fatty Amphipathic and fatty acid tail at Most abundant acid/acyl oxygen 1 & 2) Kind and Phosphate Sphingosine proportions vary backbone (fatty acid tail among membranes 3 oxygens at oxygen 2) Glycolipid Sugar head Glycerol Form by the Sphingosine addition of carbohydrates to lipids Most common are cerebrosides (uncharged sugar as head group) Prominent in brain and nerve cells Most located in the outer membrane layer of animal cells amphipathic Sterol 4 rigid & planar, Eukaryotic fused ring membranes contain structure significant amount Hydroxyl group Animals attached to ring cholesterol structure to help o Fluidity orient within buffer membrane and Plantphytosterols bond to polar Fungalergosterol head amphipathic The movement of lipids from one monolayer to another (transverse diffusion) is rare o Occur when phospholipid translocators or flippases are present Membrane fluidity changes with temperature (Tm transition temperature) Lipids move within their monolayer o Rotation o Lateral diffusion Membrane Proteins PROTEIN WHERE IDENTIFIERS EXAMPLES Integral Embedded in Transmembrane Protein Difficult to Membrane the lipid remove/isolate Cross once (singlepass Proteins bilayer b/c of from membrane proteins) hydrophobic Integral Cross several times (multipass region proteins) monotropic proteins are Anchored to the lipid bilayer embedded in one by one or more hydrophobic layer transmembrane segments Transmembrane Example: B barrel proteins span the Singlepass Membrane Protein membrane and C & Nterminus on either end protrude on both of the membrane sides Glycophorin Detergents pull Multipass Membrane Protein apart the 220 transmembrane segments membrane and bacteriohodopsin surround the protein o Micelles prevent the protein from breaking down Peripheral Hydrophilic Easy to remove regulatory protein subunits of many Proteins and located from the ion channels and transmembrane on the surface membrane receptors of the bilayer, Mostly attached hydrophilic but loosely to the some head hydrophobic regions anchor them to the membrane Electrostatic forces and hydrogen bonding (weak) Easy to isolate by altering pH or ionic strength Chelating agents are used to solubilize them Lipid Hydrophilic Reside on Fatty acidanchored membrane Anchored and attached membrane protein Proteins to the bilayer surface Attached to a saturated fatty by covalent Linked to fatty acid, usually myristic (14C) or attachments acids (isoprenyl palmitic acid (16C) to lipid groups) Isoprenylated membrane proteins molecules Synthesized in the cytosol & that are modified by addition of embedded in multiple isoprenyl groups the bilayer (5C) usually farnesy (15C) or geranylgeranyl (20C) groups GPIanchored membrane proteins Covalently linked to gylcosylphosphatidylinositol Detergents are used to disrupt hydrophobic interactions and dissolve the lipid bilayer Proteins can be solubilized and extracted from membranes so that they can be studied o Ionic Detergents Sodium deoxycholate Sodium dodecylsulfate (SDS) Attach/bind around protein Break hydrogen bonds and straighten the protein out o Nonionic Detergents Triton X100 Octylglucoside Hello! Type of Transport Important Concepts Stuff it includes Simple (passive) Solute moves from Gases Diffusion high to low Nonpolar molecules concentration Small polar molecules Moved down (water, glycerol, ethanol) concentration gradient Nothing bigger than 0.28nm No metabolic energy Increase polarity=decrease required hydrophobicity=decrease Membrane protein not permeability required Increase size=decrease Driven by permeability concentration gradient Partition coefficient K (measures No competitive hydrophobicity) the bigger the inhibition number the easier it passes through the ALWAYS moves bilayer solutes toward equilibrium Tends toward decreasing free energy Facilitated (mediated) Moves down Small polar (H2O, glycerol) Diffusion concentration gradient Large polar (glucose) No metabolic energy Ions (NA+, K+, Ca2+) required Ion Channels Membrane proteins 1. Gated are required a. Voltagechanges in Has competitive membrane potential inhibition 2. Ligand a. Triggered by certain Reversible GLUT1 substances binding Uniporter to the channel protein Transport glucose through the membrane 3. Mechanosensitive by alternating conformation a. Respond to mechanism mechanical forces acting on the Highly selective membrane T1 (conformational state) has the binding site open for glucose Porins outside of the cell Selective hole in a T2 has the binding site membrane open to the inside of Formed a multipass the cell transmembrane protein Hexokinase B barrel immediately adds a phosphate to the Aquaporin glucose to prevent to Porin is just big enough for water to pass through from leave the cell though GLUT1 Gates on the porin contain Direction on transport o Highly conserved is dictated by the Arg and His relative solute residues concentrations in and o 2 Asparagine out of cell (glucose residues who’s side concentration is kept chains form low inside most hydrogen bonds animal cells) with imported water molecules alpha helix Active Transport Not reversible Ptype Members of a large family Up concentration gradient and are reversibly Membrane protein and phosphorylated by ATP on metabolic energy a specific aspartic acid required residue 3 Indirect Inhibited by vandate VO 4 Depends on the P1 found in all organisms simultaneous transport and transport heavy metal of 2 solutes ions Favorable movement P2 responsible for of one solute down its maintaining gradients of gradient drives the ions (Na+, K+, H+, Ca2+) unfavorable across plasma membranes of eukaryotic cells, plays a movement of the other up its gradients role in muscle contractions Symport (two and acidification of gastric molecules in same juices direction, or Antiport P3 (plants/fungi) pump protons out across the (two molecules in plasma membrane, different directions) acidifying the external Driven by medium concentration gradient P4 pump hydrophobic (NOT ATP molecules (cholesterol/fatty HYDROLYSIS) acids), they do not transport SGLTNa+ follows them all the way across the concentration gradient biayer but act as flippases back into the cell and P5 not well characterized, glucose tags along but some are known the Anion Exchange transport cations Vtype Protein Facilitates reciprocal Pumps protons into exchange of Cl and organelles such as vacuoles, HCO 3 vesicles, lysosomes, endosomes, and the golgi Exchange will stop if either anion is absent complex, plasma membrane Strict 1:1 ratio of osteoclast, kidney tubules Have two multisubunit CO 2 n blood cell is components integral converted to HCO Direct component embedded in the Transport system membrane and peripheral component that’s just coupled to an outside the membrane exergonic chemical reaction, most surface commonly the Ftype Found in bacteria, hydrolysis of ATP mitochondria and Unidirectional/has an intrinsic directionality chloroplasts Na+/K+ Two components a transmembrane pore (F )0 Transport Proteins and a peripheral membrane driven by ATP hydrolysis are called component (F 1 that transport ATPases or contains the ATPbinding site ATPase pumps ALSO FUNCTION IN 3 important cellular functions REVERSEmore accurately o Uptake of called ATP synthase (not only can ATP be used as an essential energy source to generate nutrients ion gradients, but gradients o Removal of wastes can be used as an energy source to synthesize ATP) o Maintenance ABCtype of Have 4 protein domains, nonequilibriu two which are highly m hydrophobic and embedded concentrations in the membrane of certain ions The other 2 domains are peripheral and associated with the cytoplasmic side of the membrane, involved with ATP binding Pump antibiotics or drugs out of cells, rendering the cell resistant to the drug – MDR transport protein MDR transports a wide range of chemically dissimilar drugs Na+/K+ Most Proteins and their steps covered in the table o Glut 1: This is the uniporter that carries glucose into/out of the blood cell. Integral protein with 12 transmembrane segments that forms a cavity with hydrophilic side chains glucose concentration is kept low on the inside of the cell at most times How it works: 1. T1 conformation is open to the outside of cell 2. Glucose comes and binds to GLUT 1 3. The binding of glucose cause the protein to change conformation to the inside of the cell 4. The glucose is released which cause the cell to conform back into T1 This porter is reversible o This is done by glucose binding to the protein from the inside and the protein will transport it back out o So if glucose is transported into the cell why does it not rebind to the protein and cause a conformational shift back out? This is because the glucose is immediately reacted with hexokinase and phosphorylated to yield glucose 6 phosphate. Since the proteins are specific the molecule cannot renter the cytoplasm o Glucose uptake requires energy Despite glucose being indirectly brought across the membrane, it still needs a steep gradient of sodium to do so This is made possible by the Na/ K pump. Therefore these proteins are called sodiumdependent glucose transporters (SGLT) Steps of the Symporter SGLT: o 2 Na atoms bind to the protein from the outside o Glucose then binds o Conformational change from outside to inside o 2 Na release from protein (locks symporter) (they are then forced back out of the cell via Na/ K pump) glucose will release and allow the protein to change back Anion exchange protein for Cl in and HCO3 out is an antiporter o If an anion is not present the protein will stop o 1:1 ratio o The waste CO2 from the tissue enters the erythrocyte via diffusion and then it is converted into carbonate by carbonic anhydrase (this builds up one side of the gradient o As it is pushed out along its gradient, the Cl is brought in o “Ping Pong” The Chloride binds to one side (serves across the membrane) where the bicarbonate binds and (comes right back across) Na/K pump (PType) o At rest POTASSIUM IS HIGER ON THE OUTSIDE OF THE CELL AND SODIUM IS HIGHER ON THE INSIDE o 2 Conformations o E1 open to the INSIDE = (3 sodium at a time )affinity for Na o E2 open to the outside= (2 Potassium at a time) affinity for potassium o Responsible for asymmetric distribution o Contains 3 subunits, alpha beta and gamma o alpha is on the cytoplasmic side and binds ATP and Na o Steps: o In E1 3 Na bind o triggers ATP to bind o E1E2 o 3 Na release and now it is K’s turn o Bind of K triggers the dephosphorylating of the alpha subunit o E2E1 o K releases Nucleus Overview Attached to the rough ER Site where chromosomes are localized and replicated and the DNA they contained is transcribed Bounded by a nuclear envelope with an inner and an outer membrane space separated by a perinuclear space The outer membrane is continuous with the ER and contains proteins that bind actin and intermediate filaments (Ifs) of the cytoskeleton Nucleus Overview Attached to the rough ER Site where chromosomes are localized and replicated and the DNA they contained is transcribed Bounded by a nuclear envelope with an inner and an outer membrane space separated by a perinuclear space The outer membrane is continuous with the ER and contains proteins that bind actin and intermediate filaments (Ifs) of the cytoskeleton Nuclearlocalization signal (NLS) Enable the protein to be recognized and transported to the nuclear pore complex 830 amino acids in length Has 7 positive basic residues o Mechanically integrate with the rest of the cell If you change 1 amino acid you will change the destination of the protein Near the Cterminus (many protein locators on the Nterminus) Nuclear Matrix (nucleoskeleton)insoluble fibrous network that helps maintain the shape of the nucleus Nuclear Lamina thin dense meshwork of fibers lining the inner surface of the inner nuclear membrane Made on intermediate filaments made from lamins Extra layer of protection Chromatin Located in the nucleus in a nonrandom fashion Chromatin fibers extend and disperse throughout the nucleus Always arranged in the same order Nucleolus Functions in assembling ribosomal subunits in eukaryotes No nucleolus=no life There are NO ACTIVE ribosomes in the nucleolus Protein targeting All proteins translated in the cytosol Folded proteins are transported through nuclear pore complexes Large proteins/RNA Some proteins are too large to easily diffuse through the nuclear pore Large particles are actively transported across the membrane Nuclear Pores Specialized channels in the nuclear envelope where inner and outer membranes are fused Provide direct contact between the cytosol and the nucleoplasm Lined with a protein structure called Nuclear Pore Complex (NPC) o Center granule is called the transporter and is likely involved in moving molecules across the nuclear envelope Enzymes must be imported from the cytoplasm RNA’s that need to be translated and components of ribosomes must be exported from the nucleus Simple diffusion of small molecules through nuclear pores The NPC contains tiny aqueous diffusion channels through which small particles freely move Importing and Exporting in the Nucleus Nuclear Import via Importin/Ran dependent o Steps: Importin: Binds to the NLS on the protein and mediates it way into the nucleus Transported into the nucleus via the importinprotein complex The importin brings it into the nucleus and releases the protein and bind to Ran Then the RanGTP complex goes back out of the cell through the NPC Then importin dissociates by the hydrolysis of GTP Repeat Nuclear Import Ran independent o Mediated by Ca o Not much to know here Nuclear Export: o Similar to import o Used for RNA molecules o Can be Ran independent or Dependent o You have Nuclear Export Signals (NES) o Target proteins and the bound RNA for export o NES recognized by exportins o Dependent: Exportin binds to Cargo at the EPS The exportincargo complexes binds to RanGTP complex They are transported out of the cell via the NPC by the NTF2( Nuclear Transport Factor) GTP is hydrolyzed via GAP and the cargo dissociates as well as the rest of the complex. Exportin and RanGDP come back across the membrane RanGDPRanGTP via the enzyme GEF o Independent: NO RAN mRNA use this pathway Steps: o mRNa is bound by NXF1 and NX1 (these are chaperon proteins that prevent the mRNA from wadding up o Transportation through the nucleus o Rna helicase (Dbp5) comes and removes the chaperon proteins by hydrolyzing ATP o NXF1/NXT1 come back across the membrane using the RanDependent import process Nuclear Export Mainly used for RNA molecules Some traffic (mRNA) doesn’t seem to appear Ran RNA export is mediated by adaptor proteins that bind to the RNA Adaptor proteins contain sequences called nuclear export signals (NES)target proteins and the bound RNAs for export NES sequences are recognized by exportins which mediate transport of the complexes out of the nucleus Maintaining a RanGTP Gradient across the membrane Need to maintain a constant gradient RanGTP is maintained at high levels inside the nucleus by a guaninenucleotide exchange factor (GEF) that promotes Ran to bind GTP The cytosol contains a GTPase activating protein (GAP) that promotes hydrolysis of GTP by Ran The high nuclear RanGTP promotes the release of NLScontaining cargo from importin Import into ER mRNA is sent to a free ribosome to be translated Once the ER signal sequences comes out, it is bound by the SRP or (signal recognition particle) to the hydrophobic region o SRP Made up of 2 Subunits FFA or p54 FtsY or RP recα o GTP bound in protein o Hydrophobic binding groove binds to hydrophobic binding region The binding of the SRP causes translation to stop and thus it moves towards the ER o This is called DOCKING THE TRANSLOCON The SRP binds the stopped protein complex to a SRP receptor in with it is attached to a translocon by hydrolyzing 1 GTP and the SRP dissociates and translation can continue through the translocon o Translocon has a SRP and Ribosome Receptor o It also has a pore protein that forms a channel to allow the polypeptide to enter the ER and signal peptidase removes the ER signal sequence The channel opens GTP is hydrolyzed again and the SRP releases The protein folds in the Lumen of the ER A word about puromycin: o Completely stops all translation with the ER o This completely wipes out micro flora in the gut o Makes you extremely susceptible to Cdiff This produces a toxin that binds to your villi Types of ER proteins Type 1 Put in membrane o ER recognition sequence o SRPbrings it to the ER o Stop transfer sequence is translated and the rest of the protein can’t go through o peptidyl transferase cuts off signal sequence. o Nterminus goes in first translocon will put it in the membrane Type 2 (backwards) Flip type 1 o N terminus is on the cytosolic side Never had a signal sequence on that side Signal anchor seq is what got it down to the translocon Therefore, Cterm goes in first Type 3 o No signal o The signal anchor is close to Nterminus o Similar to Type 1! Be Careful Tail Anchored o The signal is on Cterminus goes into Get3 o protein complex recog and brings to ER (binds get 1+2) o Moves CEnd into lipid bilayer by hydrolysis of ATP Type 4 N terminus in exoplasmic space (Multipass) o Various stop transfer sequences cause the multiple passes Even # Cterminus on the same side as the Nterminus Odd # Nterminus and Cterminus on opposite ends GPIAnchored Proteins o The GPI molecule has both a polar (Nterminus end which contains carbohydrate and phosphate residues) and nonpolar ends (Cterminus which contains fatty acyl tails) o GPI transmidase cleaves the nonpolar end which is in the membrane and only allows into to interact on one leaflet rather than both. And transfers the carboxyl group of the polar half of an amino group on the bilayer. o Bam GPIanchored
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