Module 7 Worksheet

Phospholipid biosynthesis at the interface between the endoplasmic reticulum (ER) and the cytosol presents a number of challenges that must be solved by the cell. Explain how each of the following is handled. A) The substrates for phospholipid biosynthesis are all water soluble, yet the end products are not. B) The immediate site of incorporation of all newly synthesized phospholipids is the cytosolic leaflet of the ER membrane, yet phospholipids must be incorporated into both leaflets. C) Many membrane systems in the cell, such as the plasma membrane, are unable to synthesize their own phospholipids, yet these membranes must also expand if the cell is to grow and divide.

a. Membrane phospholipids are synthesized at the interface between the cytosolic leafl et of the endoplasmic reticulum (ER) and the cytosol. Watersoluble, small molecules are synthesized and activated in the cytosol. Membrane-bound enzymes of the ER then link these small molecules to create larger, hydrophobic membrane phospholipids. b. Membrane phospholipids can be flipped from the cytosolic leaflet of the ER membrane to the exoplasmic leaflet. This process, mediated by flippases, results in the incorporation of newly synthesized phospholipids into both leaflets. c. Phospholipids can be moved from their site of synthesis to other membranes (e.g., to the plasma membrane). Some of this transport is by vesicles. Some is due to direct contact between membranes. Small, soluble lipid-transfer proteins also mediate transfer. The mechanism of phospholipid transfer between membranes is not yet well understood.

What is the likely identity of these membrane-associated proteins: (a) a protein that is released from a membrane treated with a high-salt solution, which causes disruption of ionic linkages; (b) a protein that is not released from the membrane upon its exposure to a high-salt solution alone, but is released when the membrane is incubated with an enzyme that cleaves phosphate-glycerol bonds and covalent linkages are disrupted; (c) a protein that is not released from the membrane upon exposure to a high-salt solution, but is released after the addition of the detergent sodium dodecylsulfate (SDS). Will the activity of the protein released in part (c) be preserved following its release?

a. peripheral b. lipid anchored c. integral membrane protein. No, a strong ionic detergent like SDS will denature the protein.

Lipid rafts: A) are more fluid than the surrounding membrane. B) are less fluid than the surrounding membrane. C) are able to flip from inside to outside. D) detach from the plasma membrane and clog arteries. E) have higher rates of lateral diffusion of lipids and proteins.

are less fluid than the surrounding membrane

Intestinal transepithelial glucose transport uses a symport to transport glucose up its concentration gradient via cotransport of

glucose with Na+

Following the production of membrane extracts using the non-ionic detergent Triton X-100, you analyze the membrane lysates via mass spectrometry and note a high content of cholesterol and sphingolipids. Furthermore, biochemical analysis of the lysates reveals potential kinase activity. What have you probably isolated?

lipid raft

Cells have different types of receptors on their surface. After receptor-mediated endocytosis, where would you expect to find these receptor molecules?

on the inside surface of the vesicle

A hydropathy plot of a protein serves to:

predict whether a given protein sequence contains membrane-spanning segments.

Which of these molecules pass through a cell membrane most easily? A) large and hydrophobic B) small and hydrophobic C) large and polar D) small and charged E) none of the above

small and hydrophobic

When membrane bilayers are frozen and then fractured, they tend to split along the middle thus separating the two leaflets. This may be because:

the hydrophobic interactions that hold the membrane together are weakest at this point

Membrane depolarization can cause the opening of _______________ channels.

voltage-gated

Identify the following membrane-associated proteins based on their structure: (a) tetramers of identical subunits, each with six membrane-spanning α helices; (b) trimers of identical subunits, each with 16 β sheets forming a barrel-like structure.

(a) aquaporins (b) porins

Phospholipid biosynthesis at the interface between the ER and the cytosol presents a number of challenges that must be solved by the cell. Explain how each of the following is handled: 1.) The substrates for phospholipid biosynthesis are all water soluble, and yet the end products are not. 2.) The immediate site of incorporation of all newly synthesized phospholipids is the cytosolic leaflet of the ER membrane, and yet the phospholipids must be incorporated into both leaflets. 3.) Many membrane systems in the cell--e.g., the plasma membrane--are unable to synthesize their own phoshpolipids, and yet these membranes must expand if the cell is to grow and divide.

1.) The substrates for phospholipid biosynthesis are all water soluble because they are synthesized at the interface between the ER and the cytosol, an area where the production of hydrophobic molecules (like the end products) would not be favorable because of the hydrophilic environment at the interface. Thus, water soluble, hydrophilic substrates for phospholipid biosynthesis are synthesized in the cytosol. From there, the enzymes from the ER connect these substrates to end up creating the hydrophobic, phospholipid end products. 2.) This challenge is handled by flippases. That is, newly synthesized phospholipids in the cytosolic leaflet of the ER membrane can be flipped and then incorporated to the exoplasmic leaflet with the help of flippases. 3.) Even though membrane systems like the plasma membrane are unable to synthesize their own phospholipids, they can still expand by incorporating phospholipids that have been synthesized elsewhere. More specifically, vesicles can transport phospholipids to the membrane system, some phospholipids can be transferred from other membranes, and lipid transfer proteins can also help disperse phospholipids.

Which of these is true for the action of Na+/K+ ATPase on the plasma membranes of cells? A) 2 Na+ out, 3 K+ in, coupled to hydrolysis of 1 ATP to ADP + Pi. B) 3 Na+ out, 2 K+ in, coupled to hydrolysis of 1 ATP to ADP + Pi. C) 3 Na+ in, 2 K+ out, coupled to hydrolysis of 1 ATP to ADP + Pi. D) 1 Na+ out, 1 K+ in, coupled to hydrolysis of 1 ATP to ADP + Pi. E) 2 Na+ out, 3 K+ in, coupled to conversion of 1 ADP + Pi to ATP.

3 Na+ out, 2 K+ in, coupled to hydrolysis of 1 ATP to ADP + Pi.

Describe a negative feedback mechanism for controlling a rising cytosolic Ca2+ concentration in cells that require rapid changes in Ca2+ concentration for normal functioning. How would a drug that inhibits calmodulin activity affect cytosolic Ca2+ concentration regulation by this mechanism? What would be the effect on the function of, for example, a skeletal muscle cell?

A rise in cytosolic Ca2+ concentration causes activation of calmodulin. Some Ca2+-ATPase pumps are activated by Ca2+-calmodulin, which lowers the cytosolic Ca2+ concentration by pumping Ca2+ either into the sarcoplasmic reticulum/endoplasmic reticulum or out of the cell. An anti-calmodulin drug would inhibit this negative feedback mechanism, leaving higher Ca2+ concentration in the cytosol for a longer period of time. In skeletal muscle cells, the result would be to prolong the length and/or strength of muscle contraction.

Proteins may be bound to the exoplasmic or cytosolic face of the plasma membrane by way of covalently attached lipids. What are the three types of lipid anchors responsible for tethering proteins to the plasma-membrane bilayer? Which type is used by cell-surface proteins that face the external medium? By glycosylated proteoglycans?

Cytosolic proteins are anchored to the plasma membrane by acylation or prenylation. In the case of acylation, an N-terminal glycine residue of a protein is covalently attached to the 14-carbon fatty acid myristate (myristoylation) or a cysteine residue in a protein is attached to the 16-carbon fatty acid palmitate (palmitoylation). Prenylation occurs when the -SH group on a cysteine residue at or near the C-terminus of the protein is bound through a thioether bond to either a farnesyl or a geranylgeranyl (prenyl) group. Cell-surface proteins and heavily glycosylated proteoglycans are present on the exoplasmic face of the membrane and are linked there by a glycophosphatidylinositol (GPI) anchor

Glucose- H+ symporter is a proton based secondary active transport mechanism that brings glucose into a cell. Which of these treatments would increase the rate of glucose transport into the cells? A) Decrease extracellular glucose concentration B) Decrease extracellular pH C) Decrease cytoplasmic pH D) Adding an inhibitor that blocks the regeneration of ATP E) Adding a substance that makes the membranes more permeable to protons

Decrease extracellular pH

Glucose is a six-carbon sugar, and ribose is a five-carbon sugar. Despite its smaller size, ribose is not efficiently transported by GLUT1. How can this be explained?

Despite its smaller size, ribose cannot bind to GLUT1 as glucose does because it cannot form the same noncovalent bonds—and thus cannot be transported.

What are detergents? How do ionic and non-ionic detergents differ in their ability to disrupt biomembrane structure?

Detergents are amphipathic molecules. The hydrophobic part of a detergent molecule readily interacts with the hydrophobic tails of the phospholipids disrupting their interaction with each other; the hydrophilic part readily associates with water. This breaks up the organization of the lipid bilayer, ultimately leading to formation of micelle droplets, composed of a single phospholipid layer with the polar heads in contact with water and a hydrophobic core excluding water. Ionic detergents, like all detergents, bind to both the hydrophilic and hydrophobic regions of membrane proteins that have been exposed after lipid bilayer disruption. Because of their charge, they can also disrupt the ionic and hydrogen bonds holding together the secondary and tertiary structure of a protein and are thus useful for completely denaturing a protein. Non-ionic detergents do not denature proteins and are therefore useful for extracting membrane proteins while maintaining their native conformation. At concentrations below the critical micelle concentration, they also prevent the hydrophobic regions of proteins that have been extracted from the cell membrane from interacting with each other and forming insoluble aggregates.

Design a set of experiments to prove that GLUT1 is indeed a glucose-specific uniporter rather than a galactose- or mannose-specific uniporter.

Determination of the rate of erythrocyte (GLUT1) transport of substrate versus concentration allows the determination of Km for glucose versus galactose versus mannose. The Km for glucose will be lowest, indicating that GLUT1 is glucose-specific, not galactose- or mannose-specific.

Explain why the coupled reaction ATP → ADP + Pi in the P-class ion pump mechanism does not involve direct hydrolysis of the phosphoanhydride bond.

Direct hydrolysis of the phosphoanhydride bond would result in release of the bond energy as heat, which would thus be "lost." By fi rst transferring the phosphate bond to an aspartate (D) residue, the P-class ATPase uses the released bond energy to drive a conformational change in the protein from the E1 to the E2 state.

Patch clamping can be used to measure the conductance properties of individual ion channels. Describe how patch clamping can be used to determine whether or not the gene coding for a putative K+ channel actually codes for a K+ or a Na+ channel.

Expression of a channel protein in a normally nonexpressing cell permits the patch clamp assessment of channel properties. Typically, the cell used is a frog oocyte. Frog oocytes do not normally express plasma membrane channel proteins. Channel protein expression may be induced by microinjection of in vitro-transcribed mRNA encoding the protein. Frog oocytes are large and hence technically easier to inject and to patch clamp than other cells. One can then vary the ionic composition of the medium and determine whether the presence of Na+ or of K+ supports ionic movement through the channel.

Describe how FRAP is used to measure the movement of lipids and proteins within the bilayer. When examined by FRAP, certain integral membrane proteins are significantly less mobile than others. What accounts for this reduced mobility?

FRAP (fluorescence recovery after photobleaching) is a technique where lipids and proteins are fluorescently tagged so that the lateral movement of lipids and proteins within the bilayer can be observed. More specifically, fluorescent reagents bind to lipids while antigen-binding sites of proteins are marked with a fluorescent dye. After the initial fluorescent labeling of proteins and lipids, an area on the cell membrane is bleached which then reduces the fluorescence in that particular area. After this, we are then able to measure the movement of lipids and proteins within the bilayer by observing the non-bleached, fluorescently labeled lipids and proteins that diffuse into the bleached area. Ultimately this allows us to see how many lipids and proteins are mobile within the membrane. When examined by FRAP, certain integral membrane proteins do not diffuse into bleached area as readily which means they are less mobile than others. This is due to the fact that some integral membrane proteins, which have three domains (cytosolic, exoplasmic, and intermembrane area) and go across the phospholipid bilayer, that contain membrane-spanning α-helical domains tend to be more integrated and thus less mobile within the bilayer. Attachment to ECM or cytoskeletal proteins contributes to their reduced fluidity. In other words, it is more energetically favorable for integral membrane proteins with membrane-spanning α-helices to remain in place (not move laterally as often) due to hydrophobic and van der Waals and ionic interactions of the side-chains of the α-helices and phospholipids in the membrane.

Discuss the different factors that affect the degree of membrane fluidity.

Factors like the abundance of cholesterol, saturation and length of fatty acid chains, and overall temperature affect the degree of membrane fluidity. The presence of cholesterol determines whether or not the membrane will maintain its fluidity (by increasing it) when temperature drops or decreasing fluidity when temperatures are higher. If there is an abundance of saturated fatty acid chains, the membrane will be less fluid, but if there is an abundance of unsaturated fatty acid chain, the membrane will be more fluid. If there are more short fatty acid chains, less "packing" occurs, and they membrane will become more fluid at lower temperatures. If there are more long fatty acid chains, more "packing" occurs, and the membrane will become more fluid only at higher temperatures.

Fatty acids must associate with lipid chaperones in order to move within the cell. Why are these chaperones needed, and what is the name given to a group of proteins that are responsible for this intracellular trafficking of fatty acids? What is the key distinguishing feature of these proteins that allows fatty acids to move within the cell?

Fatty acids have very low solubility inside an aqueous-rich intracellular environment. Therefore, they associate with fatty-acid binding proteins (FABPs), which are cytosolic proteins that contain a hydrophobic pocket or barrel, lined by beta sheets. This pocket provides a haven for the long chain fatty acid, where it interacts in a noncovalent fashion with the FABP.

Describe the symport process by which cells lining the small intestine import glucose. What ion is responsible for the transport, and what two particular features facilitate the energetically favored movement of this ion across the plasma membrane?

Glucose uptake from the intestinal lumen into the epithelial cells is driven by symport with 2 Na+ ions by a 2 Na+/glucose symporter. Binding of two Na+ ions and one glucose molecule to high-affinity, outward-facing sites in the protein causes a series of conformational changes in the symporter that eventually allows Na+ and glucose to be released from low-affi nity sites facing the cytosol. Transport by this symporter is energetically favorable because movement of Na+ ions into the cell is driven by both its concentration gradient and the transmembrane voltage gradient. Transport of two Na+ ions into the cell provides ~6 kcal of energy—enough to generate an intracellular glucose concentration that is 30,000 times higher than in the intestinal lumen.

The enzyme that catalyzes the critical regulatory step of the cholesterol biosynthesis pathway is

HMG CoA reductase

What is the purpose of having cholesterol in mammalian plasma membranes?

It enables the membrane to stay fluid more easily when temperature drops.

When the proteins of the Na+/K+ ATPase are first synthesized on the ER membrane, what side of this membrane will the ATP-binding site be on?

It will be on the cytoplasmic side of the ER.

Nitric oxide (NO) is a gaseous molecule with lipid solubility similar to that of O2 and CO2. Endothelial cells lining arteries use NO to signal surrounding smooth muscle cells to relax, thereby increasing blood flow. What mechanism or mechanisms would transport NO from where it is produced in the cytoplasm of an endothelial cell into the cytoplasm of a smooth muscle cell, where it acts?

Like O2 and CO2, NO passively diffuses through membranes. As it is produced by an enzyme and accumulates in the endothelial cell cytosol, NO passively diffuses down its concentration gradient though the endothelial cell plasma membrane out of the cell and then passively diffuses through the plasma membrane into the cytoplasm of the smooth muscle cell, where it acts to decrease contraction.

Lipid bilayers are said to behave like two-dimensional fluids. What does this mean? What drives the movement of lipid molecules and proteins within the bilayer? How can such movement be measured? What factors affect the degree of membrane fluidity?

Lipid bilayers are considered to be two-dimensional fluids because lipid molecules (and proteins if present) are able to rotate along their long axes and move laterally within each leaflet. Such movements are driven by thermal energy, and may be quantified by measuring fluorescence recovery after photobleaching, the FRAP technique. In this technique, specific membrane lipids or proteins are labeled with a fluorescent reagent, and then a laser is used to irreversibly bleach a small area of the membrane surface. The extent and rate at which fluorescence recovers in the bleached area, as fluorescent molecules diffuse back into the bleach zone and bleached molecules diffuse outward, can be measured. The extent of recovery is proportional to the fraction of labeled molecules that are mobile, and the rate of recovery is used to calculate a diffusion coefficient, which is a measure of the molecule's rate of diffusion within the bilayer. The degree of fluidity depends on factors such as temperature, the length and saturation of the fatty acid chain portion of phospholipids, and the presence/absence of specific lipids such as cholesterol.

How do liver and muscle cells maximize glucose uptake without changing Vmax?

Liver cells convert glucose to glycogen, which maximizes the glucose gradient across the plasma membrane.

The membrane potential in animal cells, but not in plants, depends largely on resting K+ channels. How do these channels contribute to the resting membrane potential? Why are these channels considered to be nongated channels? How do these channels achieve selectivity for K+ versus Na+, which is smaller than K+?

Membrane potential refers to the voltage gradient across a biological membrane. The generation of this voltage gradient involves three fundamental elements: a membrane to separate charge, a Na+/K+ ATPase to achieve charge separation across the membrane, and nongated K+ channels to selectively conduct current. The major ionic movement across the plasma membrane is that of K+ from inside to outside the cell. Movement of K+ outward, powered by the K+ concentration gradient generated by Na+/K+ ATPase, leaves an excess of negative charges on the inside and creates an excess of positive charges on the outside of the membrane. Thus, an inside-negative membrane potential is generated. These potassium channels are referred to as resting K+ channels. This is because these channels, although they alternate between an open and closed state, are not affected by membrane potential or by small signaling molecules. Their opening and closing are nonregulated; hence, the channels are called nongated. K+ channels achieve selectivity for K+, versus, say, Na+, through coordination of the nonhydrated ion with carbonyl groups carried by amino acids within the channel protein. The ion enters the channel as a hydrated ion, the water of hydration is exchanged for interaction with carbonyl residues within the channel, and then as the ion exits the channel it is rehydrated. Within the confines of the channel protein structure, Na+, unlike K+, is too small to replace fully the interactions of water with those with amino acid-carried carbonyl groups. Because of this, the energetic situation is highly unfavorable for Na+ versus K+.

Name the three groups into which membrane-associated proteins may be classified. Explain the mechanism by which each group associates with a biomembrane.

Membrane-associated proteins may be classifi ed as integral membrane proteins, lipid-anchored membrane proteins, or peripheral membrane proteins. Integral membrane proteins pass through the lipid bilayer and are therefore composed, of three domains: a cytosolic domain exposed on the cytosolic face of the bilayer; an exoplasmic domain exposed on the exoplasmic face of the bilayer; and a membrane-spanning domain, which passes through the bilayer and connects the cytosolic and exoplasmic domains. Lipid-anchored membrane proteins have one or more covalently attached lipid molecule, which embeds in one leaflet of the membrane and thereby anchors the protein to one face of the bilayer. Peripheral proteins associate with the lipid bilayer through interactions with either integral membrane proteins or with phospholipid heads on one face of the bilayer.

Phospholipids and cholesterol must be transported from their site of synthesis to various membrane systems within cells. One way of doing this is through vesicular transport, as is the case for many proteins in the classic secretory pathway (see Chapter 14). However, phospholipid and cholesterol membrane-to-membrane transport in cells does not occur solely by vesicular transport. What is the evidence for this statement? What appear to be the major mechanisms for phospholipid and cholesterol transport?

Most phospholipids and cholesterol membrane-to-membrane transport in cells is not by Golgi-mediated vesicular transport. One line of evidence for this is the effect of chemical and mutational inhibition of the classical secretory pathway. Either fails to prevent cholesterol or phospholipid transport between membranes, although they do disrupt the transport of proteins and Golgiderived sphingolipids. Membrane lipids produced in the ER cannot move to the mitochondria by classic secretory transport vesicles. No vesicles budding from the ER have been found to fuse with mitochondria. Other mechanisms are thought to exist. However, presently these are poorly defined. They include direct membrane-membrane contact and small, soluble lipid-transfer proteins.

Acetic acid (a weak acid with a pKa of 4.75) and ethanol (an alcohol) are each composed of two carbons, hydrogen, and oxygen, and both enter cells by passive diffusion. At pH 7, one is much more able to permeate a cellular membrane than the other. Which is more membrane permeable, and why? Predict how the membrane permeability of each is altered when the extracellular pH is reduced to 1.0, a value typical of the stomach.

Of the two at neutral pH, ethanol is the much more membrane permeant. It has no acidic or basic group and is uncharged at a pH of 7.0. The carboxyl group of acetic acid is predominantly dissociated at this pH and hence acetic acid exists predominantly as the negatively charged acetate anion. It is nonpermeant. At a pH of 1.0, ethanol remains uncharged and membrane permeant. The carboxyl group of acetic acid is now predominantly nondissociated and uncharged. Hence, acetic acid is now membrane permeant. Any difference in permeability is very small.

Which of the following is true? A) Phospholipids can laterally diffuse along the plane of the membrane. B) Phospholipids frequently flip-flop from one leaflet to the other. C) Phospholipids occur in an uninterrupted sheet, with proteins restricted to the surfaces of the membrane. D) Phospholipids are free to depart from the membrane and dissolve in the surrounding solution. E) Phospholipids have hydrophilic tails in the interior of the membrane.

Phospholipids can laterally diffuse along the plane of the membrane.

Plants use the proton electrochemical gradient across the vacuole membrane to power the accumulation of salts and sugars in the organelle. This accumulation creates hypertonic conditions in the vacuole. Why does this not result in the plant cell swelling and bursting? Even under isotonic conditions, there is a slow leakage of ions into animal cells. How does the plasma-membrane Na+/K+ ATPase enable animal cells to avoid osmotic lysis under isotonic conditions?

Plant cells, unlike animal cells, are surrounded by a cell wall. This cell wall is relatively stiff and rigid. The hyperosmotic situation within the plant vacuole that typically constitutes most of the volume of the plant cell is resisted by the rigid cell wall and the cell does not burst. Overall, a plant cell is considered to have a turgor pressure because of the hyperosmotic vacuole. The Na+/K+ ATPase is key to animal cells avoiding osmotic lysis. Animal cells have a slow inward leakage of ions. In the absence of a countervailing export, this would result in osmotic lysis of the cells even under isotonic conditions. The main countervailing export is the net transport of cations by Na+/K+ ATPase (3 Na+ ions out for 2 K+ in).

Although both faces of a biomembrane are composed of the same general types of macromolecules, principally lipids and proteins, the two faces of the bilayer are not identical. What accounts for the asymmetry between the two faces?

Since biomembranes form closed compartments, one face of the bilayer is automatically exposed to the interior of the compartment while the other is exposed to the exterior of the compartment. Each face therefore interacts with different environments and performs different functions. The different functions are in turn directly dependent on the specific molecular composition of each face. For example, different types of phospholipids and lipid-anchored membrane proteins are typically present on the two faces. In addition, different domains of integral proteins are exposed on each face of the bilayer. Finally, in the case of the plasma membrane, the lipids and proteins of the exoplasmic face are often modified with carbohydrates

Explain the mechanism by which statins lower "bad" cholesterol.

Statins block the conversion b-hydroxy-b-methylglutaryl linked to CoA (HMG-CoA) to mevalonate (an important intermediate in cholesterol synthesis) by competitively binding the enzyme necessary for this conversion (HMG-CoA reductase).

Movement of glucose from one side to the other side of the intestinal epithelium is a major example of transcellular transport. How does the Na+/K+ ATPase power the process? Why are tight junctions essential for the process? Why is localization of the transporters specifically in the apical or basolateral membrane crucial for transcellular transport? Rehydration supplements such as sport drinks include a sugar and a salt. Why are both important to rehydration?

The Na+/K+ ATPase located on the basolateral surface of intestinal epithelial cells uses energy from ATP to establish Na+ and K+ ion gradients across the intestinal epithelial cell plasma membrane. Cotransporters couple the energetically unfavorable movement of glucose and amino acids into epithelial cells to the energetically favorable movement of Na+ into these cells. The accumulation of glucose and amino acids here is an important example of secondary active transport. Tight junctions are essential for the process because they seal the interstitial space between cells and hence allow the transport proteins in the apical and basolateral membranes of the epithelial cell to be effective. Effective transport could not be achieved through a leaky cell layer. The coordinated transport of glucose and Na+ ions across the intestinal epithelium creates a transepithelial osmotic gradient. This forces the movement of water from the intestinal lumen across the cell layer and hence promotes water absorption from sport drinks.

Explain the following statement: The structure of all biomembranes depends on the chemical properties of phospholipids, whereas the function of each specific biomembrane depends on the specific proteins associated with that membrane.

The amphipathic nature of phospholipid molecules (a hydrophilic head and hydrophobic tail) allows these molecules to self-assemble into closed bilayer structures when in an aqueous environment. The phospholipid bilayer provides a barrier with selective permeability that restricts the movement of hydrophilic molecules and macromolecules across the bilayer. The different types of proteins present on the two faces of the bilayer contribute to the distinctive functions of each membrane, and control the movement of selected hydrophilic molecules and macromolecules across it.

What are the common fatty acid chains in phosphoglycerides, and why do these fatty acid chains differ in their number of carbon atoms by multiples of 2?

The common fatty-acid chains in phosphoglycerides include myristate, palmitdate, stearate, oleate, linoleate, and arachidonate (see Table 2-4). These fatty acids differ in carbon atom number by multiples of 2 because they are elongated by the addition of 2 carbon units. For example, the acetyl group of acetyl CoA is a 2-carbon moiety

What is the molecular basis of potassium channel's ability to select for K+ over Na+?

The differential interaction of these cations with the selectivity filter

How do fatty acids with double bonds help keep membranes fluid when temperatures drop?

The double bonds form kinks in the fatty acid tails and prevent tight packing.

Why do aquaporins fail to transport H+ whereas some can transport glycerol?

The failure of aquaporins to transport H+ can be boiled down to hydrogen bonding within the aquaporin channel. That is, in order for a molecule like glycerol or H+ to be transported through an aquaporin, they must be able to hydrogen bond with the amino acids within the channel. Glycerol, with is hydroxyl groups, can hydrogen bond and be transported through an aquaporin whereas H+ cannot hydrogen bond with the amino acids in an aquaporin and thus cannot be transported.

Name the four classes of ATP-powered pumps that produce active transport of ions and molecules. Indicate which of these classes transport ions only and which transport primarily small organic molecules. The initial discovery of one class of these ATP-powered pumps came from studying the transport not of a natural substrate, but rather of artificial substrates used as cancer chemotherapy drugs. What do investigators now think are common examples of the natural substrates of this particular class of ATP-powered pumps?

The four classes of ATP-powered pumps are: P-class, V-class, F-class, and ABC superfamily. Only the ABC superfamily members transport small organic molecules. All other classes pump cations or protons. The initial discovery of ABC superfamily pumps came from the discovery of multidrug resistance to chemotherapy and the realization that ultimately this was due to transport proteins (i.e., ABC superfamily pumps). Today, the natural substrates of ABC superfamily pumps are thought to be small phospholipids, cholesterol, and other small molecules.

Virus X infects host cells that have R cell surface molecules. The viral nucleic acid molecules are enclosed in a protein capsid, and the protein capsid is itself contained inside an envelope consisting of a lipid bilayer membrane and viral glycoproteins. One hypothesis for viral entry into cells is that binding of viral membrane glycoproteins to R initiates fusion of the X membrane with the plasma membrane of the host, releasing the viral capsid into the cytoplasm. An alternative hypothesis is that X gains entry into the cell via receptor-mediated endocytosis, and membrane fusion occurs in the endocytotic vesicle. To test these alternative hypotheses for X entry, researchers labeled the lipids on the X membrane with a green fluorescent dye. What would be observed by live-cell fluorescence microscopy if the green fluorescent lipid dye-labeled X membrane fuses with the host cell plasma membrane?

The green fluorescent dye-labeled lipids will diffuse into the infected host cell's plasma membrane.

How does muscle contraction increase when the action of the Na+/K+ ATPase is inhibited?

The inhibition of the transporter increases [Na+ ] in the cells and therefore decreases Ca2+ export.

The biosynthesis of cholesterol is a highly regulated process. What is the key regulated enzyme in cholesterol biosynthesis? This enzyme is subject to feedback inhibition. What is feedback inhibition? How does this enzyme sense cholesterol levels in a cell?

The key regulated enzyme in cholesterol biosynthesis is HMG (b-hydroxy-bmethylglutaryl)-CoA reductase. This enzyme catalyzes the rate controlling step in cholesterol biosynthesis. The enzyme is subject to negative feedback regulation by cholesterol. In fact, the cholesterol biosynthetic pathway was the first biosynthetic pathway shown to exhibit this type of end-product regulation. As the cellular cholesterol level rises, the need to synthesize additional cholesterol goes down. The expression and enzymatic activity of HMG-CoA reductase is suppressed. HMG-CoA reductase has eight transmembrane segments and, of these, fi ve compose the sterol-sensing domain. Sterol sensing by this domain triggers the rapid, ubiquitin-dependent proteasomal degradation of HMG CoA reductase. Homologous domains are found in other proteins such as SCAP (SREBP cleavage activating protein) and Niemann-Pick C1 (NPC1) protein, which take part in cholesterol transport and regulation

What key mechanistic features results in large differences in transport rates of uniporters vs. ion channels?

The large differences in transport rates of uniporters vs. ion channels basically boils down to the mechanical complexity of each transport system. Uniporters tend to be the slower channel because the transports that they process are more complex. That is, a substrate has to bind to the uniporter, the uniporter must undergo a conformational change, and the uniporter can also only transport one molecule at a time. Ion channels on the other hand are a lot faster because they don't have substrates that bind (rather move through a high-affinity chamber), they do not undergo conformational changes, and they can transport numerous substrates (ions) at once.

What do you predict will happen when you transfer a phospholipid bilayer from water to oil?

The membrane is expected to invert its normal structure such that the polar head groups hide inside of the bilayer and the non-polar fatty acid tails face out to the oil.

In the case of the bacterial two-Na+/one-leucine symporter, what is the key distinguishing feature of the bound Na+ ions that ensures that other ions, particularly K+, do not bind?

The six oxygens in the main-chain carbonyl or side-chain carboxyl groups that bind each of the two Na+ ions in the transporter are exquisitely positioned with a geometry similar to that of the water molecules with which Na+ associates in solution. At one site, the carboxyl group of the bound leucine provides one of the coordinating oxygens. When Na+ ions bind to the oxygens, they lose their water of hydration. The increase in entropy that occurs when hydration water molecules are freed promotes Na+ ions binding at both sites. K+ ions (and water molecules themselves) are too big to bind the six oxygens in the proper geometry and so do not compete with Na+.

When viewed by electron microscopy, the lipid bilayer is often described as looking like a railroad track. Explain how the structure of the bilayer creates this image.

The spontaneous assembly of phospholipid molecules into a lipid bilayer creates a sheetlike structure that is two molecules thick. Each layer is arranged so that the polar head groups of the phospholipids are exposed to the aqueous environment on one side of the bilayer and the hydrocarbon tails associate with the tails of the other layer to create a hydrophobic core. In cross section, the bilayer structure thus consists of a hydrophobic core bordered by polar head groups. When stained with osmium tetroxide, which binds strongly to polar head groups, and viewed in cross section, the bilayer looks like a railroad track with a light center bounded on each side by a thin dark line.

Name the three classes of membrane transport proteins. Explain which one or ones of these classes is able to move glucose and which can move bicarbonate (HCO3−) against an electrochemical gradient. In the case of bicarbonate, but not glucose, the ΔG of the transport process has two terms. What are these two terms, and why does the second not apply to glucose? Why are cotransporters often referred to as examples of secondary active transport?

The three classes of transporters are uniporters, symporters, and antiporters. Both symporters and antiporters are capable of moving organic molecules against an electrochemical gradient by coupling an energetically unfavorable movement to the energetically favorable movement of a small inorganic ion. The ΔG for bicarbonate has two terms, a concentration term and an electrical term, because bicarbonate is an anion. Glucose is neutrally charged and hence its ΔG for transport has only a concentration term. Unlike pumps, neither symporters nor antiporters hydrolyze ATP or any other molecule during transport. Hence, these cotransporters are better referred to as examples of secondary active transporters rather than as actual active transporters. The term active transporter is restricted to the ATP pumps where ATP is hydrolyzed in the transport process.

Compare and contrast between the three main types of lipids found in biomembranes.

The three main types of lipids found in biomembranes are steroids (cholesterol), sphingolipids, and phosphoglycerides. While all three are similar in that they have both hydrophobic and hydrophilic regions (amphipathic, polar head and hydrophobic tail), they differ from each other in "chemical structure, abundance, and function." (Lodish et al). Phosphoglycerides are unique in that they are the most abundant, have a hydrophobic tail with two fatty acid chains, a hydrophilic head connected to a phosphate group, and function as signaling molecules. Sphingolipids are unique in that they are not as abundant as phosphoglycerides, have a hydrophobic tail with one fatty acid chain which is connected to hydrophilic head (which may or may not contain a phosphate group) via an amide link, and have a protective structural function. Cholesterol (steroids) is unique in that it is most abundant in plasma membranes, has a four-ring structure, and functions as a manager of membrane fluidity.

Certain proton pump inhibitors that inhibit secretion of stomach acid are among the most widely sold drugs in the world today. What pump does this type of drug inhibit, and where is this pump located?

These drugs irreversibly inhibit the H+/K+ ATPase in the apical membrane of stomach parietal cells. Although the inhibition of a given H+/K+ ATPase is irreversible, the cells eventually make more of the pump.

An H+ ion is smaller than an H2O molecule, and a glycerol molecule, a three-carbon alcohol, is much larger. Both readily dissolve in H2O. Why do aquaporins fail to transport H+ whereas some can transport glycerol?

To be transported, a molecule must fi t into the aquaporin channel and form hydrogen bonds with N-H groups of amino acids lining the channel. Although H+ is smaller than H2O, it cannot form the required hydrogen bonds. Glycerol is much larger than H2O, but the three-carbon chain is flexible and the three OH groups can form the required hydrogen bonds.

Tumor cells expressing GLUT1 often have a higher Vmax for glucose transport than do normal cells of the same type. How could these cells increase the Vmax?

Tumor cells often express a higher number of glucose transporters than normal cells.

Uniporters and ion channels support facilitated transport across cellular membranes. Although both are examples of facilitated transport, the rates of ion movement via an ion channel are roughly 104- to 105-fold faster than the rates of molecule movement via a uniporter. What key mechanistic difference results in this large difference in transport rate? What contribution to free energy (ΔG) determines the direction of transport?

Uniporters are slower than channels because they mediate a more complicated process. The transported substrate both binds to the uniporter and elicits a conformational change in the transporter. A uniporter transports one substrate molecule at a time. In contrast, channel proteins form a protein-lined passageway through which multiple water molecules or ions move simultaneously, single file, at a rapid rate. The major contributor to the free-energy-driving transport through a uniporter is the entropy increase as a molecule moves from a high concentration to a low concentration

Why are water-soluble substances unable to freely cross the lipid bilayer of the plasma membrane? How does the cell overcome this permeability barrier?

Water-soluble substances are hydrophilic; they are therefore repelled by thehydrophobic core of the bilayer, which is composed of non-polar hydrocarbon tails of the phospholipids. Proteins that span the cell membrane (transmembrane proteins) provide a channel or passageway through which these substances can cross the membrane. The proteins fold such that their non polar residues are in contact with the phospholipid bilayer and their polar residues line the channel through which the hydrophilic substances travel from one side of the cell membrane to the other.

Fat and muscle cells modulate the Vmax for glucose uptake in response to insulin signaling. How?

When insulin is low, GLUT4 is stored in intracellular vesicles. Insulin induces a rapid increase in the Vmax for glucose uptake by stimulating fusion of these vesicles with the plasma membrane, thereby increasing the number of plasma membrane glucose transporters.

An integral membrane protein can be extracted from a membrane with:

a solution containing a detergent