How do stains used for light microscopy compare with those used for electron microscopy?
Stains used for light microscopy are colored molecules that bind to cell components, affecting the light passing through, while stains used for electron microscopy involve heavy metals that affect the beams of electrons passing through.
Which type of microscope would you use to study
a) the changes in shape of a living white blood cell
b) the details of surface texture of a hair
c) the detailed structure of an organelle
a) light microscope
b) scanning electron microscope
c) transmission electron microscope
Briefly describe the structure and function of the nucleus, the mitochondrion, the chloroplast, and the endoplasmic reticulum.
Nucleus: Nuclear envelope: is a double membrane enclosing the nucleus; perforated by pores; continuous with ER. Nucleolus: structure involved in production of ribosomes; a nucleus has one or more nucleoli. Chromatin: material consisting of DNA and proteins; visible as individual chromosomes in a dividing cell
Mitochondrion: organelle where cellular respiration occurs and most ATP is generated; double-bound membrane; folded inner membrane; has some individual DNA
Chloroplast: photosynthetic organelle; converts energy of sunlight to chemical energy stored in sugar molecules; double bound membrane; inner thylakoids; individual DNA
ER: network of membranous sacs and tubes; active in membrane synthesis and other synthetic ad metabolic processes; has rough (ribosomal studded) and smooth regions
Imagine an elongated cell (such as a nerve cell) that is 125 x 1 x 1, using arbitrary units similar to the ones in Figure 6.8. Predict where the surface-to-volume ratio would lie in Figure 6.8. Then calculate and check your prediction
This cell would have the same volume as the cells in columns 2 and 3 but proportionally more surface area than in column 2 and less than in column 3. Thus, the surface-to-volume ratio should be greater than 1.2 but less than 6. To obtain the surface area, you'd have to add he area of the six sides 125 +125 +125 +125+1+1 = 502. The surface-to-volume ratio equals 502 divided by a volume of 125, or 4.0
What role do ribosomes play in carrying our genetic instructions?
Ribosomes in the cytoplasm translate the genetic message, carried form the DNA in the nucleus by mRNA, into a polypeptide chain.
Describe the molecular composition of nucleoli and explain their function.
Nucleoli consist of DNA and the ribosomal RNA (rRNA) made according to its instructions, as well as proteins imported from the cytoplasm. Together, the rRNA and proteins are assembled into large and small ribosomal subunits. (These are exported through nuclear pores to the cytoplasm, where they will participate in polypeptide synthesis.)
If the function of a particular protein is a eukaryotic cell is to make up a part of the chromatin, describe the process of its synthesis. Include the cellular locations of all relevant molecules.
The information in a gene (on a chromosome in the nucleus) is used to synthesize an mRNA that is then transported through a nuclear pore to the cytoplasm. There it is translated into protein, which is transported back through a nuclear pore into the nucleus, where is joins other proteins and DNA, forming chromatin.
Describe the structural and functional distinctions between rough and smooth ER.
The primary distinction between rough and smooth ER is the presence of bound ribosomes on the rough ER. While both types of ER make phospholipids, membrane proteins and secretory proteins are all produced on the ribosomes of the rough ER. The smooth ER also functions in detoxification, carbohydrate metabolism, and storage of calcium ions.
Descrie how transport vesicles integrate the endomembrane system.
Transport vesicles move membranes and substances they enclose between other components of the endomembrane system.
Imagine a protein that functions in the ER but requires modification in the Golgi apparatus before it can achieve that function. Describe the protein's path through the cell, starting with the mRNA molecule that specifies the protein.
The mRNA is synthesized in the nucleus and then passes out through a nuclear pore to be translated on a bound ribosome, attached to the rought ER. The protein is synthesized into the lumen of the ER and perhaps modified there. A transport vesicle carries the protein to the Golgi apparatus. After further modification in the Golgi, another transport vesicle carries it back to the ER, where it will perform its cellular function.
Describe two common characteristics of chloroplasts and mitochondria. Consider both function and membrane structure.
Both organelles are involved in energy transformation, mitchondria in cellular respiration and chloroplasts in photosynthesis. They both have multiple membranes that separate their interiors into compartments. In both organelles, the innermost membranes - cristae, or infoldings of the inner membrane, in mitochondria, and the thylakoid membranes in chloroplasts - have large surface areas with embedded enzymes that carry out their main functions.
A classmate proposes that mitochondria, chloroplasts, and peroxisomes should be classified in the endomembrane system. Argue against the proposal.
Mitochondria, chloroplasts, and peroxisomes are not derived form the ER, nor are they connected physically or via transport vesicles to organelles of the endomembrane system. Mitochondria and chloroplasts are structurally quite different from vesicles derived from the ER, which are bounded by a single membrane.
Describe shared features of microtubule-based motion of flagella and microfilament-based muscle contraction.
Both systems of movement involve long filaments that are moved in relation to each other by motor proteins that grip, release, and grip again adjacent polymers.
How do cilia and flagella bend?
Dynein arms, powered by ATP, move neighboring doublets of microtubules relative to one another. Because they are anchored within the organelle with respect to each other, the doublets bend instead of sliding past each other.
Males afflicted with Kartagener's syndrome are sterile because of immotile sperm, tend to suffer lung infections, and frequently have internal organs, such as the heart, on the wrong side of he body. This disorder has a genetic basis. Suggest what the underlying defect might be.
Such individuals have defects in the microtubule-based movement of cilia and flagella. Thus, the sperm can't move because of malfunctioning flagella; the airways are compromised; signaling events during embryogenesis do not occur directly due to malfunctioning cilia.
In what way are the cells of plants and animals structurally different from single-celled eukaryotes?
The most obvious difference is the presence of direct cytoplasmic connections between cells of plants (plasmodesmata) and animals (gap junctions). These connections result in the cytoplasm being continuous between adjacent cells.
IF the plant cell wall or the animal extracellular matrix were impermeable, what effect would this have on cell function?
The cell would not be able to function properly and would probably soon die, as the cell wall or ECM must be permeable to allow the exchange of matter between the cell and its external environment. Molecules involved with energy production and use must be allowed entry, as well as those that provide information about the cell's environment. Other molecules, such as products synthesized by the cell for export and the by-products of cellular respiration, must be allowed to exit.
Which statement correctly characterizes bound ribosomes?
Bound ribosomes generally synthesize membrane proteins and secretory proteins.
Which structure is NOT part of the endoplasmic system?
Cells of the pancreas will incorporate radioactively labeled amino acids into proteins. This "tagging" of newly synthesized proteins enables a researcher to track their location. In this case, we are tracking an enzyme secreted by pancreatic cells. What is its most likely pathway?
ER-->Golgi-->vesicles that fuse with plasma membrane
Which structure is common to plant AND animal cells
Which of the following is present in a prokaryotic cell?
Which cell would be best for studying lysosomes?
phagocytic white blood cell
Which structure-function pair is mismatched?
microtubule; muscle contraction
Cyanide binds with at least one molecule involved in producing ATP. If a cell is exposed to cyanide, most of the cyanide would be found within the
The carbohydrates attached to some proteins and lipids of the plasma membrane are added as the membrane is made and refined in the ER and Golgi apparatus; the new membrane then forms transport vesicles that travel to the cell surface. On which side of the vesicle membrane are the carbohydrates?
They are on the inner side of the transport vesicle membrane.
How would you expect the saturation levels of membrane phospholipid fatty acids to differ in plants adapted to cold environments and plants adapted to hot environments?
Plants adapted to cold environments would be expected to have more unsaturated fatty acids in their membranes because those remain fluid at lower temperatures. Plants adapted to hot environments would be expected to have more saturated fatty acids, which would allow the fatty acids to "stack" more closely, making the membranes less fluid and therefore helping them to stay intact at higher temperatures.
Two molecules that can cross a lipid bilayer without help from membrane proteins are O2 and CO2. What properties allow this to occur?
O2 and CO2 are both small nonpolar molecules that can easily pass through the hydrophobic core of a membrane.
Why would water molecules need a transport protein to move rapidly and in a large quantities across a membrane?
Water is a polar molecule, so it cannot pass very rapidly through the hydrophobic region in the middle of the phosopholipid bilayer.
Aquoporins exclude passage of hydronium ions (H3O+). But recent research has revealed a role for some aquoporins in fat metabolism, in which they allow passage of glycerol, a three-carbon alcohol as well as H2O. Since H3O+ is much closer in size to water than is glycerol, what do you suppose is the basis of this selectivity?
The hydronium ion is charged, while glycerol is not. Charge is probably more significant than size as a basis for exclusion by the aquaporin channel.
How do you think a cell performing cellular respiration rids itself of the resulting CO2?
CO2 is a small nonpolar molecule that can diffuse through the plasma membrane. As long as it diffuses away so the concentration remains low outside the cell, it will continue to exit the cell in this way. (This is the opposite of the case for O2.)
In the supermarket, produce is often sprayed with water. Explain why this makes vegetables look crisp.
The water is hypotonic to the plant cells, so they take up water and the cells of the vegetable remain turgid, rather than plasmolyzing. The vegetable, (for example, lettuce or spinach) remains crisp and not wilted.
If a Paramecium swims from a hypotonic environment to an isotonic one? will its contractile vacuole become more active or less? Why?
The activity of the Paramecium's contractile vacuole will decrease. The vacuole pumps out excess water that flows into the cell; this flow occurs only in a hypotonic environment.
When nerve cells establish a voltage across their membrane with a sodium-potassium pump, does this pump use ATP or does it produce ATP? Why?
The pump uses ATP. To establish a voltage, ions have to be pumped against their gradients, which requires energy.
Explain why the sodium-potassium pump would not be considered a cotransporter.
Each ion is being transported against its electrochemical gradient. If either ion were flowing down its electrochemical gradient, this WOULD be considered cotransport.
What would happen if cells had a channel protein allowing unregulated passage of hydrogen ions?
Even if proton pumps were still using ATP and moving protons, no proton gradient would become established. This would have serious consequences for the cells, because processes like the cotransport of sucrose (as well as synthesis of ATP) depend on establishment of a proton gradient.
As a cell grows, its plasma membrane expands. Does this involve endocytosis or exocytosis? Explain.
Exocytosis. When a transport vesicle fuses with the plasma membrane, the vesicle membrane becomes part of the plasma membrane.
To send a signal, a neuron may carry out exocytosis of signaling molecules that are recognized by a second neuron. In some cases, the first neuron ends the signal by taking up the molecules by endocytosis. Would you expect this to occur by pinocytosis or by receptor-mediated endocytosis? Explain.
Receptor-mediated endocytosis. In this case, one specific kind of molecule needs to be taken up at a particular time; pinocytosis takes up substances in the nonspecific manner.
In what way do the membranes of a eukaryotic cell vary?
Certain proteins are unique to each membrane.
According to the fluid mosaic model of membrane structure, proteins of the membrane are mostly
embedded in a lipid bilayer
Which of the following factors would tend to increase membrane fluidity?
A greater proportion of unsaturated phospholipids
Which of the following processes includes all others?
b) diffusion of a solute across a membrane
c) facilitated diffusion
d) passive transport
e) transport of an ion down it electrochemical gradient
d) passive transport
Which of these experimental treatments would increase the rate of sucrose transport into the cell?
decreasing extracellular pH
How does the second law of thermodynamics help explain the diffusion of a substance across a membrane?
The second law is the trend toward randomness. Equal concentrations of a substance on both sides of a membrane is a more random distribution than unequal concentrations. Diffusion of a substance to a region where it is initially less concentrated increases entropy, as described by the second law.
Describe the forms of energy found in an apple as it grows on a tree, then falls ans is digested by someone who eats it.
The apple has potential energy in its position hanging on the tree, and the sugars and other nutrients it contains have chemical energy. The apple has kinetic energy as it falls from the tree to the ground. Finally, when the apple is digested and its molecules broken down, some of the chemical energy is used to do work, and the rest is lost as thermal energy.
If you place a teaspoon of sugar in the bottom of a glass of water, it will dissolve completely over time. Left longer, eventually the water will disappear and the sugar crystals will reappear. Explain these observations in terms of entropy.
The sugar crystals become less ordered (entropy increases) as they dissolve and become randomly spread out in the water. Over time, the water evaporates, and the crystals form again because the water volume is insufficient to keep them in solution. While the reappearance of sugar crystals may represent a "spontaneous" increase in order (decrease in entropy), it is balanced by the decrease in order (increase in entropy) of the water molecules, which changed from a relatively compact arrangement in liquid water to a much more dispersed and disordered form in water vapor.
Cellular respiration uses glucose and oxygen, which have high levels of free energy, and releases CO2 and water, which have low levels of free energy. Is respiration spontaneous or not? Is it exergonic or endergonic? What happens to the energy released from glucose?
Cellular respiration is a spontaneous and exergonic process. The energy released from glucose is used to do work in the cell or is lost as heat.
A key process in metabolism is the transport of hydrogen ions (H+) across a membrane to create a concentration gradient. Other processes can result in an equal concentration of hydrogen ions on each side. Which arrangement of hydrogen ions allows the H+ to perform work in this system?
Hydrogen ions can perform work only if their concentrations on each side of a membrane differ. When the H+ concentrations are the same, the system is at equilibrium and can do no work.
At nighttime celebrations, revelers can sometimes be seen wearing glow-in-the-dark necklaces. The necklaces start glowing once they are "activated," which usually involved snapping the necklace in a way that allows two chemicals to react and emit light inthe form of "chemiluminescence." Is the chemical reaction exergonic or endergonic? Explain your answer.
The reaction is exergonic because it releases energy - in this case, in the form of light.
In most cases, how does ATP transfer energy from exergonic to endergonic reactions in the cell?
ATP transfers energy to endergonic processes by phosphorylating (adding phosphate groups) to other molecules. (Exergonic processes phosphorylate ADP to regenerate ATP.)
What of the following combinations has more free energy?
a) glutamic acid + ammonia + ATP
b) glutamine + ADP + P
Explain your answer.
A set of coupled reactions can transform the first combination into the second. Since, overall, this is an exergonic process, ∆G is negative and the first group must have more free energy.
Many spontaneous reactions occur very slowly. Why don't all spontaneous reactions occur instantly?
A spontaneous reaction is a reaction that is exergonic. However, if it has a high activation energy that is rarely attained, the rate of the reaction may be low.
Why do enzymes act only on very specific substrates?
Only the specific substrate(s) will fit properly into the active site of an enzyme, the part of the enzyje taht carries out catalysis.
Malonate is an inhibitor of the enzyme sucinate deydrogenase. Howwould you determine whether malonate is a competitive or noncompetitive inhibitor?
Increase the concentration of the normal substrate (succinate) and see whether the rate of reaction increases. If it does, malonate is a competitive inhibitor.
How can an activator and an inhibitor have different effects of an allosterically regulated enzyme?
The activator binds in such a way that it stabilizes the active form of an enzyme, whereas the inhibitor stabilizes the inactive form.
Imagine you are a pharmacological researcher who wants to design a drug that inhibits a particular enzyme. Upon reading the scientific literature, you find that the enzyme's active site is similar to that of several other enzymes. What might be the best approach to developing your inhibitor drug?
You might choose to screen chemical compounds that bind allosterically to the enzyme, because allosteric regulatory sites are less likely to share similarity between different enzymes.
Choose the pair of terms that correctly completes this sentence:
Catabolism is to anabolism as ___________ is to __________.
Most cells cannot harness heat to perform work because
temperature is usually uniform throughout a cell
Which of the following metabolic processes can occur without a net influx of energy from some other process?
C6 H12 O6 + 6O2 --> 6CO2 + 6H2O
In an enzyme in solution is saturated with substrate, the most effective way to obtain a faster yield of products is to
add more of the enzyme
If an enzyme is added to a solution where its substrate and products are in equilibrium, what would occur?
Nothing; the reaction would stay at equilibrium
Some bacteria are metabolically active in hot springs because
their enzymes have high optimal temperatures
L can form either M or N
M can form O
O can form either P or R
P can form Q
R can form S
O inhibits the reaction of L to form M
Q inhibits the reaction of O to form P
S inhibits the reaction of O to form R
Which reaction would prevail if both Q and S were present in the cell in high concentrations?
Compare and contrast aerobic and anaerobic respiration.
Both process include glycolysis, the citric acid cycle, and oxidative phophorylation. In aerobic respiration, the final electron acceptor is molecular oxygen(O2), whereas in anaerobic respiration, the final electron acceptor is a different substance.
If the following redox reaction occurred, which compound would be oxidized and which reduced?
C4H6O5 + NAD+ --> C4H4O5 +NADH + H+
C4H6O5 would be oxidized and NAD+ would be reduced.
During the redox reaction in glycolysis, which molecule acts as the oxidizing agent? The reducing agent?
NAD+ acts as the oxidizing agent, accepting elecrons from glyceraldehyde-3-phosphate, which thus acts as the reducing agent.
Step 3 in Figure 9.9 is a major point of regulation of glycolysis. The enzyme phosphofructokinase is allosterically regulated by ATP and related molecules. Considering the overall result of glycolysis, would you expect ATP to inhibit or stimulate activity of this enzyme? (Hint: Make sure you consider the role of ATP as an allosteric regulator, not as a substrate of he enzyme.)
Since the overall process of glycolysis results in net production of ATP, it would make sense for the process to slow down when ATP levels have increased substantially. Thus we would expect ATP to allosterically inhibit phosphofructokinase.
Name the molecules that conserve most of the energy from the citric acid cycle's redox reactions. How is this energy converted to a form that can be used to make ATP?
NADH and FADH2; they will donate electrons to the electron transport chain.
What cellular processes produce the CO2 that you exhale?
CO2 is released from the pyruvate that is formed during glycolysis, and CO2 is also released during the citric acid cycle.
The conversions shown in Figure 9.10 and step 4 of Figure 9.12 are each catalyzed by a large multienzyme complex. What similarities are there in the reactions that occur in these two cases?
In both cases, the precursor molecule loses a CO2 molecule and then donates electrons to an electron carrier in an oxidation step. Also, the product has been activated due to the attachment of a CoA group.
What effect would an absence of O2 have on the process shown in Figure 9.16?
Oxidative phosphorylation would stop entirely, resulting in no ATP production by this process. Without oxygen to "pull electrons down the electron transport chain, H+ would not be pumped into the mitochondrion's intermenbrane space and chemiosmosis would not occur.
In the absence of O2, as in the question above, what do you think would happen if you decreased the pH of the intermembrane space of the mitochondrion? Explain your answer.
Decreasing the pH is the addition of H+. It would establish a proton gradient even without the function of the electron transport chain, and we would expect ATP synthase to function and synthesize ATP. (In fact, it was experiments like this that provided support for chemiosmosis as an energy-coupling mechanism.)
Consider the NADH formed during glycolysis. What is the final acceptor for its electrons during fermentation? What is the final acceptor of its electrons during aerobic respiration?
A derivative of pyruvate - such as actaldehyde during alcohol fermentation - or pyruvate itself during lactic acid fermentation; oxygen.
A glucose-fed yeast cell is moved from an aerobic environment to an anaerobic one. For the cell to continue generating ATP at the same rate, how would its rate of glucose consumption need to change?
The cell would need to consume glucose at a rate about 19 times the consumption rate in the aerobic environment (2 ATP are generated by fermentation versus up to 38 ATP by cellular respiration).
Compare the structure of a fat with that of a carbohydrate. Wat features of their structures make a fat a much better fuel?
The fat is much more reduced; it has many - CH2 - units, and in all these bonds the electrons are equally shared. The electrons present in a carbohydrate molecule are already somewhat oxidized (shared unequally in bonds), as quite a few of them are bound to oxygen.
Under what circumstances might your body synthesize fat molecules?
When we consume more food than necessary for metabolic processes, our body synthesizes fat as a way of storing energy for later use.
What will happen in a muscle cell that has used up its supply of oxygen and ATP?
AMP will accumulate, stimulating phosphofructokinase, which increases the rate of glycolysis. Since oxygen is not present, the cell will convert pyruvate to lactate in lactic acid fermentation, providing a supply of ATP.
What is the reducing agent in the following reaction?
Pyruvate + NADH + H+ --> Lactate + NAD+
The IMMEDIATE energy source that drives ATP synthesis by ATP synthase during oxidative phosphorylation is the
H+ concentration across the membrane holding ATP synthase
Which metabolic pathway is common to both fermentation and cellular respiration of a glucose molecule?
In mitochondria, exergonic redox reactions
provide the energy that establishes the proton gradient
The final electron acceptor of the electron transport chain that functions in aerobic oxidative phosphorylation is
When electrons flow along the electron transport chains of mitochondria, which of the following changes occurs?
the pH of the matrix increases
Cells do not catabolize carbon dioxide because
CO2 is already completely oxidized
Which of the following is a true distinction between fermentation and cellular respiration?
NADH is oxidized by the electron transport chain in respiration only
Most CO2 from catabolism is released during
the citric acid cycle
How do the reactant molecules of photosynthesis reach the chloroplasts in leaves?
CO2 enters leaves via stomata, and water enters via roots and is carried to leaves through veins.
How did the use of an oxygen isotope help elucidate the chemistry of photosynthesis?
Using 18O, a heavy isotope of oxygen, as a label, van Niel was able to show that the oxygen produced during photosynthesis originates in water, not in carbon dioxide.
The Calvin cycle clearly requires the products of the light reactions, ATP and NADPH. Suppose a classmate asserts that the converse is not true - that the light reactions don't depend on the Calvin cycle, and, with continual light, could just keep on producing ATP and NADPH. Do you agree or disagree? Explain.
The light reactions could NOT keep producing NADPH and ATP without the NADP+, ADP, and P that the Calvin cycle generates. The two cycles are interdependent.
What color of light is LEAST effective in driving photosynthesis? Explain.
Green, because green light is mostly transmitted and reflected - not absorbed - by phtosynthetic pigments.
Compared to a solution of isolated chlorophyll, why do intact chloroplasts release less heat and fluorescence when illuminated?
In chloroplasts, light-excited electrons are trapped by a primary electron acceptor, which prevents them from dropping back to the ground state. In isolated chlorophyll, there is no electron acceptor, so the photoexcited electrons immediately drop back down to the ground state, with the emission of light and heat.
In the light reactions, what is the initial electron donor? Where do the electrons end up?
Water (H2O) is the initial electron donor; NADP+ accepts electrons at the end of the electron transport chain, becoming reduced to NADPH.
In an experiment, isolated chloroplasts placed in a solution with the appropriate components can carry out ATP synthesis. Predict what would happen to the rate of synthesis if a compound is added to the solution that makes membranes freely permeable to hydrogen ions.
In this experiment, the rate of ATP synthesis would sow and eventually stop. Because the added compound would not allow a proton gradient to build up across the membrane, ATP synthase could not catalyze ATP production.
To synthesize one glucose molecule, the Calvin cycle uses ________ molecules of CO2, ________ molecules of ATP, and _________ molecules of NADPH.
6; 18; 12
Explain why the large numbers of ATP and NADPH molecules used during the Calvin cycle are consistent with the high value of glucose as an energy source.
The more potential energy a molecule stores, the more energy and reducing power is required for the formation of that molecule. Glucose is a valuable energy source because it is highly reduced, storing lots of potential energy in its electrons. To reduce CO2 to glucose, much energy and reducing power are required int he form of large numbers of ATP and NADPH molecules, respectively.
Explain why a poison that inhibits an enzyme of the Calvin cycle will also inhibit the light reactions.
The light reactions require ADP and NADP+, which would not be formed in sufficient quantities from ATP and NADPH if the Calvin cycle stopped.
Explain why photorespiration lowers photosynthetic output for plants.
Photorespiration decreases photosynthetic output by adding oxygen, instead of carbon dioxide, to the Calvin cycle. As a result, no sugar is generated (no carbon is fixed), and O2 is used rather than generated.
The presence of only PS I, not PS II, in the bundle-sheath cells of C4 plants has an effect on O2 concentration. What is that effect, and how might that benefit the plant?
Without PS II, no O2 is generated in bundle-sheath cells. This avoids the problem of O2 competing with CO2 for binding to rubisco in these cells.
How would you expect the relative abundance of C3 versus C4 and CAM species to change in a geographic region whose climate becomes much hotter and drier?
C4 and CAM species wold replace many of the C3 species.
The light reactions of photosynthesis supply the Calvin cycle with
ATP and NADPH
Which of the following sequences correctly presents the flow of electrons during photosynthesis?
H2O --> NADPH --> Calvin cycle
In mechanism, photophosyphorylation is most similar to
oxidative phosphorylation in cellular respiration
How is photosynthesis similar in C4 plants and CAM plants?
In both cases, rubisco is not used to fix carbon initially.
Which process is most directly driven by light energy?
removal of electrons form chlorophyll molecules
Which of the following statements is a correct distinction between autotrophs and heterotrophs?
Autotrophs, but not heterotrophs, can nourish themselves beginning with CO2 and other nutrients that are inorganic.
Which of the following does NOT occur during the Calvin cycle?
release of oxygen