A nucleosome consists of a segment of DNA wrapped around a disc shaped complex of proteins called histones. These histone proteins bind to the phosphate backbone of the DNA. Describe the types of molecular bonds that would be necessary to assemble a nucle
amino acid bonds, hydrogen bonding interactions between nucleotides, hydrophobic/hydrophilic interactions, cysteine....DNA backbone is negatively charged: electrostatic interaction...(cysteine bonds); DNA consists of two long polymers of simple units called nucleotides, with backbones made of sugars and phosphate groups joined by ester bonds (to carboxyl of amino acid) ; histones make five types of interactions with DNA: * Helix-dipoles from alpha-helices in H2B, H3, and H4 cause a net positive charge to accumulate at the point of interaction with negatively charged phosphate groups on DNA * Hydrogen bonds between the DNA backbone and the amide group on the main chain of histone proteins * Nonpolar interactions between the histone and deoxyribose sugars on DNA * Salt links and hydrogen bonds between side chains of basic amino acids (especially lysine and arginine) and phosphate oxygens on DNA * Non-specific minor groove insertions of the H3 and H2B N-terminal tails into two minor grooves each on the DNA molecule
Account for the 36 ATPs produced pper molecule of glucose oxidized during cellular respiration (including glycolysis). Be ast detailed as you can in your answer. Be sure to include the subcellular locations of the different portions of the process.
Because oxygen was not present in the atmosphere until plants began to produce it through photosynthesis, early organisms needed to find a way to get the energy out of food with out the use of oxygen. This process 1s sometimes called fermentation but is more correctly called glycolysis. Glycolysis begins with sugar, usually glucose. The activation energy for this reaction is 2 ATPs. The glucose molecule (a 6 carbon compound) is broken down to two three carbon compounds. The bonds of this three carbon compound are rearranged and 2 ATPs worth of energy are gained from each molecule and 1 molecule of NADH. The new molecule is called pyruvic acid. If oxygen were present, the hydrogen on the NADH would be converted to water and more ATPs would be produced. Because there is no oxygen present the hydrogen atom is passed on to another three carbon compound, called 2 lactic acid or a two carbon compound called ethyl alcohol and a molecule of CO2. It depends on the organism which of these two products is formed. It terms of energy, the cell lost 2 ATPs to start the reaction and only gained 4 ATPs (2 from each formation of pyruvic acid). There is energy in NADH, but there is no oxygen so there is no way to get it out. This is a very small gain in energy which is why only prokaryotes or single celled creatures can survive on glycolysis alone. This process occurs in the cytoplasm of the cell because this process was developed before mitochondria evolved. The next process takes the end product of glycolysis, pyruvic acid, and is able to remove the remaining energy from the bonds. This process is called the citric acid cycle or the Krebs cycle. (Citric acid because it is involved in the process, Krebs because he was the man who figured it out). The Citric acid cycle occurs in the mitochondria and begins by converting pyruvic acid into acetate, a two carbon compound. This process releases a CO2 molecule and forms a molecule of NADH. The acetate joins with an enzyme carrier called Coenzyme A and forms a compound called Acetyl CoA. This goes through a complex series of chemical reactions which forms 1 ATP molecule, three NADH molecules, 1 FADH2 molecule, (the FADH2 molecule is another energy-carrying molecule) and two CO2 molecules. Since NADH and FADH2 can not be used by the cell directly for energy, they need to go through another process called the electron transport system. This "cashes in" the energy-carrying molecules to ATP. Each NADH can produce 3 ATPs and each FADH2 can produce 2 ATP molecules. This occurs when the hydrogen is passed from compound to another, releasing a little bit of energy each time. The final hydrogen acceptor is oxygen, forming water. This is why we need oxygen to live, to provide the final hydrogen acceptor in the electron transport system. The energy count for the entire process is as follows: 2 ATPs for Glycolysis 2 ATP 2 NADH for Glycolysis 6 ATP 2 NADH for pyruvic acid to acetate 6 ATP 2 ATPs from Citric Acid Cycle 2 ATP 6 NADH from Citric Acid Cycle 18 ATP 2 FADH2 from Citric Acid Cycle 4 ATP TOTAL = 38 ATP But the activation energy for this reaction was 2 ATPs so the net gain by the cell is 36 ATPs
Autoradiography and/or GFP fusion proteins can be used to track the flow of proteins through the endomembrane system. What are the final destinations of proteins that are processed in this pathway? Also, can you think of a technique that involves breaking
In the secretory pathway, proteins move from the ER to the golgi and can be observed using these two techniques, which showed that they eventually end up in an organelle, the plasma membrane, or the extracellular space. Subcellular fractionation is a complimentary technique, which can help to determine the molecular composition by breaking microsomes up into smooth and rough fractions and then specific enzymes can be isolated and then antipodies can be made for it and used to visualize it later with electron microscopy.
Calculate delta G for a the movement of amole of Na+ ions OUT of a cell with a -70mV membrane potential, T=25degrees celsius and an outside Na+ concentration of 150mM and an internal concentration of 15mM. What does the sign of your answer tell you?
DeltaG=2.303RTlog[Na(out)]/[Na(in)] - zFdeltam; t = 298k, z = +1, m = -.07V, F = 23.06kCal/V(M) = delta G = 2.303(.00198kcal/mol(K))298KLog(0.15M/.015M) - (1)23.06kcal/v-m(-.07V);;;;;;;;;;;Delta G = 2.303 RT log ( [Co]/[Ci] ) -Z F Delta Esubm R = 1.987 cal/mol K T= 273 + 25 = 298 K Delta Esubm = -70mV *Clore's notes: best to convert to kcal Answer = 2.97 kcal/mol = 2970 cal/mol
Describe how translation is initiated and terminated (do not need to describe elongation)
Translation is initiated when a small ribosomal subunit and initiation factor proteins join to form a 43S complex. This complex then scans the mRNA for the start codon AUG. When it is found, the complex binds to the codon and the initiation factors are released. The large ribosomal subunit then binds to the small subunit and codon and initiation is complete. Termination occurs when the process encounters a termination codon UUA, UGA, or UAG. When the encountered, a release factor is brought in instead of another codon, and this covalently bonds to the peptide strand. Doing so also causes the hydrolysis of GTP, which causes the peptide to be released (Clore's notes: "via ribosome in large subunit") from the ribosome and the subunits to disengage from each other and the RNA.
Describe some of the techniques that have been used to determinet that proteins are mobile within the lipid bilayer. How might some of these same techniques be used to test for indications of the presence of lipid rafts?
used to think it was solid...then lipid bilayer...then through freeze fracture see the proteins, SEM...then fluid mosaic model in the 1960's...flourescense recovery after photobleaching (FRAP) -- a component of the membrane is labeled with a floursecent dye...a small region is irradiated to bleach the dye, and teh recovery of flourescense in the bleached region is followed...single particle tracking (SPT) -- individual membrain proteins are labled (usually with antibody-coated gold particles) and the movements of the proteins are followed by commputer enhanced video microscopy...fractioniation of membrane proteins using polyacrylamide gel electrophoresis (PAGE) with red blood cells...evidence collected from 1970-1990 showed evidence for the existence of aggregations of different aggregations of different lipids into microdomains within membranes...specifically cholesterol and sphingolipids These can be found by applying detergent...The GPI proteins have resistance to detergent...during centrifugation they will become separated.
Describe the microarray technique (the basics of both how and why it is performed, as well as strengths and weaknesses)
to monitor the expression of thousands of genes expressed in a particular cell population...DNA fragments representing individual genes are generated using PCR, DNA cloning. The cloned DNAs are spotted in an array on a slide. mRNAs present in the cells are purified, converted to a population of flourescently labeled complementary DNAs. The DNAs that have been hybridized are identified by examination of the slide under the microscope. Any spot in the microarray that exhibits flourescense represents a gene that has been trascribed in the cells being studied....expensive....(fast though)...use of computer decreases error rate...don't need large samples
Describe the regulated changes in a fibroblast's actin cytoskeleton that allow the cell to migrate across a surface. Mention the roles of specific actin binding proteins whenever possible.
(??)The cytoplasmic extensions reach forward, bind to the substrate, the majority of the cell pulls itself forward, and the reat extensions separate from the substrate. In terms of proteins and the actin cytoskeleton, it is more complicated. The leading edge of the extensions move due to an actin actin nucleating complex known as ARP 2/3, which is activated by a complex known as WASP. Myosin II and unconventional Myosin I are believe to be involved in the process when the cell pulls itself forward to the leading edge. The release of rear extensions from the substrate occurs when the ARP 2/3 complex is broken down. These proteins and complexes, as well as the actin filaments themselves, are carried and moved by the motor protein myosin.
Discuss the reasons why Rubisco might be considered a flawed enzyme, but also why it is fundamental to life.
The slow turnover rate (approx 3.2-3.3 s-1) compared to other enzymes (104-105 s-1). Lack of specificity. Where it is supposed to fix CO2 it can and does with only a little less affinity to fix O2 instead. When it does this, the light independent rxn have to release already fixed CO2, which costs the cell the 02, CAM & C4 plants have ways to reduce rubisco. Oxygen fixing is call photorespiration.
Draw a diagram of a mitochondrion with all compartments and membranes labeled and containing ATP synthases (indicate the direction of H+ flow through them) and the electron transport chain (and indicating the directionality of H+ pumping). Would you expec
http://en.wikipedia.org/wiki/Mitochondria....yes...would change medium pH...in order to maintain proton motive force, through the use of uncopling proteins...outer membrane is hihgly permeable
Explain how kinesin was discovered. Would this same basic strategy work for retrieving myosin from extracts? Why or why not?
kinesin was discovered in 1985 from the cytoplasm of squid giant%2
Explain why the following is true or false: Individual proteins are RNA molecules are able to enter and exit the nucleus through the nuclear pores by themselves (without the aid of other molecules) through the openings formed by the "spokes" of the pores
The nuclear pore complex consists of a spoke ring assembly, a nuclear basket, and eight cytoplasmic filaments....small soluble solutes could go through...larger things like proteins/RNA cannot get through unassisted...need nuclear localization signal...p490...probably know basic steps...big proteins need exporins...for RNA need EJC (491)
Starting with a cell of a typical resting potential of -70mV and typical internal and external ion concentrations, if suddenly the plasma membrane became permeable to the same degree to both Na+ and K+, in which direction would you expect each of these io
Potassium would flow out and sodium would flow in. Initially sodium would move in at a faster rate because of the relative negative charge inside the cell. As teh concentration of sodium increased potassium K+ would begin to leave the cell rapidly through leakage channels in order to return to normal conditions. Eventually sodium potassium pumps would be activated and Na+ would be pumped back out...
Suppose you fed a culture of secretory cells non-hydrolyzable GTP (i.e. a form of GTP that would bind the relevant GTP binding proteins but could not be hydrolyzed and released). What might these cells look like under a TEM and why? You should provide as
.(probably about the microtubule assemblies...tubulin dimers, hollow cylindrical...dynamic instability due to GTP cap, allows you to build/break down the microtubules...catastrophe) -- p.314 -- without hydrolysis, not going to have this....you would see just the balls, not the (b);;;;;;;;;;;;;;;;For MT to grow, GTP is added to the dynamic/Beta/(+) polar end faster than GDP is hydrolyzed (which depolymerizes structure). Having non-hyrdolyzable GTP allows for more rapid growth in cells. GTP also causes tubules to be (more) linear and grow, whereas GDP depolymerizing catastrophe by bengin the beta end inward with a curved shape + mis-aligning/weakening the covalent bond b/w dimer strands. This facilitation of microtuble building I imagine would speed up production, also male for fat or stressed looking cells.
What are the differences between DELTA G and DELTA G Prime for a cellular reaction? Is it possible to have a positive Gprime and negativeG for the same reactoin? Why or why not? Can an enzyme change the sign of deltaG for a reaction? (can it make an ender
A) Delta G = free energy change, calculated by enthalpy - change in entropy for specific conditions/concentrations, where G delta Prime (standard free energy change) is used for standard conditions (K=298, 1ATM pressure, concentrations = I M)...leading to the equation: G = Gprime + 2.303RT logKeq B)Yes. There may be a net loss in free energy if the products formed are being broken down (as a common intermediate in a metabolic pathway e.g.) fast enough so that the concentrations of reactants is greater...C) No. It can only contribute to the rate which a reaction occurs. An enzyme does not add any energy to the reaction, it just lowers the activation requirement Ea. (all spontaneous reactions have a +G)
What are the functions of histones and how and why might they be modified? If histone H1s were removed from interphase chromatin, what would the chromatin look like and why?
Histones 1st line of DNA organization above double helix. Also can inactivate DNA, if they are methylated, which prevents gene expression. Hyper express sequence of DNA "acetylate" it rather than "methylate" if you remove H1 Histones from chromatin you would still have some attempted at increased levels of organization, so you would see something resembling what it would normally look like, but with many double helices looping in and out, beads on string.
What are the types of proteins that allow passive movement/transport of solutes across the membrane?
ion channels, controlled by special ligands like neurotransmitters, facillitative transporters, P-type active transporters, sodium + calcium ion pumps, motor proteins which may open and close them...The major types of proteins that allow passive movement are trans-membrane proteins. Types of these include channels, some of which ion channels, and gates. These methods depend mostly on concentrations of the particles going through. Receptor proteins (Clore's note: "ligand-gated channels") are also involved in some of the gated channels, opening and closing them. Diagram of channel and gate through membrane.