Questions on chloroplast structure and the light-dependent reactions

Question 1

Describe the light-dependent reactions of photosynthesis, in the order in which they happen, from the absorption of a photon to the formation of reduced NADP.

Accepted Answer

B1 a photon is absorbed by the pigments of photosystem II and the energy is funnelled to the chlorophyll a at the reaction centre, from which an electron is raised to a higher energy level and lost altogether, leaving the molecule positively charged B1 the lost electron is replaced by the photolysis of water on the inner face of the thylakoid membrane, which gives protons, electrons and oxygen B1 the excited electron is passed from carrier to carrier along the thylakoid membrane, losing energy at each transfer B1 that energy drives protons from the stroma into the thylakoid space, so a steep proton gradient builds across the membrane B1 protons flow back into the stroma through ATP synthase, driving photophosphorylation, while the electron reaches photosystem I, is re-energised by light and is passed with a proton to NADP, giving reduced NADP

Question 2

Compare non-cyclic photophosphorylation with cyclic photophosphorylation.

Accepted Answer

B1 non-cyclic uses photosystem II and photosystem I, whereas cyclic uses photosystem I alone B1 in non-cyclic the electron ends up on NADP and never returns, whereas in cyclic it is passed to a carrier and returned to the same photosystem B1 non-cyclic needs photolysis of water to replace the electrons photosystem II loses, so it releases oxygen, whereas cyclic splits no water and releases none B1 non-cyclic produces both ATP and reduced NADP, whereas cyclic produces ATP only, which suits a job such as pumping potassium ions into a guard cell

Question 3

Explain why thylakoids are stacked into grana rather than existing as single flattened sacs, and explain why the thylakoid membrane must be sealed.

Accepted Answer

B1 chlorophyll molecules are held in the thylakoid membrane, so stacking thylakoids into grana packs a large area of membrane, and so a large quantity of pigment, into a small volume B1 more membrane area means more light can be absorbed per chloroplast B1 the light-dependent stage depends on protons being concentrated on one side of the thylakoid membrane, forming a steep concentration gradient A1 a leaky membrane would let that proton gradient drain away as fast as it was built, so the membrane has to be sealed for chemiosmosis to work at all

Question 4

Describe how a chromatogram of leaf pigments is prepared, and describe why the origin line must be drawn in pencil and kept above the level of the solvent.

Accepted Answer

B1 the leaf is ground with a little propanone, and the extract is spotted onto a pencil line drawn near the bottom of a strip of chromatography paper, letting each spot dry before adding the next so it stays small and concentrated B1 the paper is then stood in a shallow layer of solvent, with the origin line held above the surface of the solvent B1 the line must be drawn in pencil because ink would dissolve in the solvent and run up the paper along with the pigments, making the result impossible to interpret A1 the origin must be kept above the solvent surface because a spot dipped directly into the solvent would dissolve straight off the paper into the solvent rather than travelling up the paper as the solvent rises

Question 5

On a chromatogram the solvent has travelled 92 mm from the origin, and the centre of the chlorophyll b spot lies 34 mm from the origin. Calculate the Rf value of chlorophyll b, and calculate how far a carotene spot would have travelled if its Rf value is 0.94 under the same conditions.

Accepted Answer

M1 Rf = distance moved by the spot ÷ distance moved by the solvent, so for chlorophyll b that is 34 ÷ 92 A1 Rf = 0.37, to two significant figures M1 for carotene, distance moved by the spot = Rf × distance moved by the solvent = 0.94 × 92 A1 86 mm, to two significant figures

Question 6

Describe how the Hill reaction, using the redox indicator DCPIP, is used to measure the rate of the light-dependent reactions in a suspension of isolated chloroplasts.

Accepted Answer

B1 DCPIP is blue when oxidised and colourless when reduced, and it is added to a suspension of isolated chloroplasts in place of NADP as an artificial electron acceptor B1 in the light, electrons excited in the chloroplasts' electron transport chain are passed to DCPIP instead of to NADP, and gaining electrons is reduction, so the DCPIP loses its blue colour B1 in the dark no electrons are excited, so DCPIP is not reduced and stays blue A1 the time taken for the blue colour to disappear, or the rate at which it fades, is a measure of the rate of the light-dependent reactions, so this can be compared between different light intensities or wavelengths

Question 7

A student varies light intensity by moving a lamp to different distances from tubes of chloroplast suspension and DCPIP, and takes the relative intensity at each distance to be 1 ÷ distance². Explain two problems with this way of varying light intensity, and explain how each can be overcome.

Accepted Answer

B1 first problem explained, one of: a lamp close to a tube warms it, and temperature changes the rate for a reason other than light; intensity is proportional to 1 ÷ distance² only for a point source, which a real lamp is not; light from the room adds an intensity that was not measured B1 the fix for that problem: a water bath at a fixed temperature with a heat filter between lamp and tubes, or an LED; a light meter reading taken where the tube stands; a darkened room with the tubes screened B1 a second, different problem from the same list, explained B1 the fix for the second problem, from the same list of fixes

Question 8

Explain why a leaf appears green, referring to the absorption spectrum of chlorophyll.

Accepted Answer

B1 chlorophyll absorbs strongly in the blue region and in the red region of the spectrum B1 its absorption falls to a small fraction of that peak in the green band around 550 nm, so green light is barely absorbed B1 the green light is reflected and transmitted instead, so it is the light that reaches the eye, and a leaf lit only by green light photosynthesises slowly

Question 9

On a chromatogram of leaf pigments the solvent front has run 84 mm from the pencil origin line, and the centre of the carotene spot lies 79 mm from that line. Calculate the Rf value of carotene.

Accepted Answer

M1 Rf = distance moved by the spot ÷ distance moved by the solvent, with both distances measured from the pencil line and the spot measured to its centre M1 79 ÷ 84 A1 Rf = 0.94, a ratio with no units, which can never exceed 1

Question 10

A plant is watered with water in which the oxygen is the heavy isotope oxygen-18, while the carbon dioxide it is given contains only ordinary oxygen. Suggest where the oxygen-18 will first be detected, and justify your answer.

Accepted Answer

B1 the oxygen-18 appears in the oxygen gas the plant releases B1 the oxygen given off in photosynthesis comes from the photolysis of water at the thylakoid membrane, not from carbon dioxide B1 water is split into protons, electrons and oxygen, so any label carried by the water leaves in the gas; had the oxygen come from carbon dioxide, unlabelled gas would have been released instead

Question 11

Explain why the presence of DNA and 70S ribosomes inside a chloroplast is used as evidence that chloroplasts evolved from free-living prokaryotic cells.

Accepted Answer

B1 DNA and 70S ribosomes are features of prokaryotic cells, and chloroplasts carry both B1 a eukaryotic cell's own ribosomes, outside its chloroplasts and mitochondria, are the larger 80S type, so a chloroplast's ribosomes do not match the rest of the cell they are found in A1 the simplest explanation for a chloroplast carrying its own DNA and a prokaryote-type ribosome is that it was once a free-living prokaryotic cell, engulfed by an ancestral eukaryotic cell and retained rather than digested

Question 12

Explain why photosystem II cannot keep absorbing light and losing electrons unless water is continuously split nearby.

Accepted Answer

B1 each time photosystem II absorbs a photon, the chlorophyll a at its reaction centre loses an electron and becomes positively charged, and it must regain an electron before it can absorb light and lose one again B1 photolysis of water, catalysed by the oxygen-evolving complex on the inner face of the thylakoid membrane, releases electrons that refill photosystem II A1 without a continuous supply of replacement electrons from water, photosystem II would quickly run out of electrons to lose and light absorption at that photosystem would stop, so the whole non-cyclic pathway depends on photolysis continuing at the same rate as photoionisation

Question 13

A poison specifically inhibits ATP synthase in the thylakoid membrane, without affecting any other protein there. Suggest what happens to the proton concentration in the thylakoid space, and suggest what happens to the rate of the light-independent reactions as a result.

Accepted Answer

B1 protons continue to be pumped into the thylakoid space by the electron transport chain, but can no longer flow back out through ATP synthase, so the proton concentration inside the thylakoid space keeps rising B1 with ATP synthase blocked, ATP production stops, even though electron transport and photolysis may continue for a while A1 the light-independent reactions need a continuous supply of ATP to reduce GP to triose phosphate and to regenerate RuBP, so with no ATP being made the light-independent reactions in the stroma slow and then stop, even though light is still available

Question 14

Compare the role of chlorophyll a with the role of the accessory pigments, chlorophyll b and the carotenoids, in capturing light energy.

Accepted Answer

B1 chlorophyll a sits at the reaction centre of every photosystem and is the only pigment able to pass an excited electron on to the next carrier, whereas the accessory pigments cannot do this themselves B1 the accessory pigments absorb wavelengths chlorophyll a absorbs poorly, particularly in the blue-green between about 450 and 520 nm, and pass the energy they capture on to chlorophyll a at the reaction centre A1 a plant with only chlorophyll a would therefore be able to use the light energy it did absorb, but would waste much of the blue-green light that the accessory pigments would otherwise capture

Question 15

In an investigation of dehydrogenase activity in chloroplasts, leaves were blended in an isolation medium that was ice-cold, buffered, and of the same water potential as the chloroplasts. Explain why each of these three properties was needed.

Accepted Answer

B1 ice-cold, so that enzymes released from the broken cells, which would digest the chloroplast membranes and proteins, work slowly B1 same water potential, so there is no net movement of water into or out of the chloroplasts by osmosis and they do not burst or shrink B1 buffered, so the pH stays constant and the proteins of the electron transport chain are not denatured

Question 16

A student follows the reduction of DCPIP by a suspension of isolated chloroplasts with a colorimeter. Explain why a red filter is used, and why the colorimeter is set to zero absorbance with a tube of chloroplast suspension and medium containing no DCPIP.

Accepted Answer

B1 oxidised DCPIP is blue, so it absorbs red light, and the absorbance of red light falls as the DCPIP is reduced and loses its colour B1 chlorophyll in the suspension also absorbs red light, and the tube without DCPIP cancels out that absorbance B1 so the reading measures only the DCPIP still oxidised, and zero absorbance is the end-point the tube reaches when all its DCPIP has been reduced

Question 17

State what an absorption spectrum measures, and state what an action spectrum measures.

Accepted Answer

B1 an absorption spectrum shows how much light a pigment absorbs at each wavelength B1 an action spectrum shows the rate of photosynthesis at each wavelength

Question 18

State the location within a chloroplast of the light-dependent reactions, and state the location of the light-independent reactions.

Accepted Answer

B1 the light-dependent reactions occur on and across the thylakoid membranes B1 the light-independent reactions occur in the stroma surrounding the thylakoids