bicd 110 wi15 problem set#1

7/24/2019 BICD 110 Wi15 Problem Set#1

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1) Extremophiles are microorganisms, usually from the Archeal domain, that grow under

a wide variety of extreme conditions (temperature, pressure, pH, salt, etc.). Some ofthese extremophiles can even grow in temperatures up to 121C, the sterilization

temperature of an autoclave. Identifying the adaptations that allow these microorganisms

to thrive at such high temperatures is an area of active research both for improving

industrial processes (i.e biofuels) and exobiology. Recent studies of the membranecomposition of found that the major lipids in these organisms are tetraether phospholipids

and diether phospholipids. In the diagram the R represents a variety of head groups.

A) Diagram what a membrane made up of diether phospholipids would look like.

Compare this to eukaryotic cell membranes.

B) Diagram what a membrane made up of tetraether phospholipids would look like.Compare this to eukaryotic cell membranes.

C) Diagram what a membrane made up of a mixture of diether and tetraether

phospholipids would look like. Compare this to eukaryotic cell membranes.

D) Would you expect the ratio of tetraether to diether phospholipids in the thermophile'smembrane to increase or decrease with increasing temperature? Give two reasons to

justify your answer.

E) Would you expect either of these phospholipids in the thermophiles's membrane to

become more or less saturated as temperature increases? Explain your answer.


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F) It turns out that thermophiles don't modify the saturation state of the lipid in response

to temperature. They do however introduce pentacyclic groups into the lipid structure inresponse to temperature (structure below). Studies have shown that increasing the

number of these groups in the tetraether phospholipids allows them to pack more tightly

together. Given this information would you expect more or fewer pentacyclic

modifications as temperature increases? Explain your reasoning.

2) If you measure the phase transition temperature (TT) of lipids isolated from E. coligrown at 37C you find that it is 22C.

A) Draw the phase transition graph marking TTand the growth temperature (TG).

B) Using your graph, do you expect the lipids to be more gel-like or more liquid-like at

the following temperatures:15C, 30C, 43C.

C) You decide to growE. coliat a range of temperatures for 24 hours, isolate the lipids,and then measure the phase transition temperature of the lipids. You obtain the following

results:

TG TT

15C 0C

30C 15C

37C 22C

43C 28C

Why does the phase transition temperature change with growth temperature? Describe 2

mechanisms that would allow this to happen.


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3) (11-34) Ion transporters are often linked together as a consequence of their actions.

For instance, cells can raise their intracellular pH by exchanging external Na+ for internalH+. The change in internal Na+ is then redressed using the Na+/K+ pump.

A) Can these two transporters working together correct the low pH without altering Na

concentration?

B)How does the linked action of these two pumps affect K+ concentration and membranepotential?

4) (11-38) A rise in intracellular Ca2+ concentration by causes muscles to contract. In

addition to an ATP driven Ca2+ pump on the sarcoplasmic reticulum (a specialized form

of ER), heart muscle cells have an antiporter on the plasma membrane that exchangesNa+ for Ca2+ as well as the standard Na+/K+ pump.

A) Draw a muscle cell with these three pumps, the direction of ion transport, whether

ATP is used, and note whether the Na+, K+, and Ca2+ concentrations are high or low.

B) Digitalis is a Na+/K+ pump inhibitor used to treat heart disease by making the heartpump more strongly. Diagram and explain how this works noting the changes in Na+,K+, and Ca2+ that occur in various compartments.

C) Digitalis in not often used clinically anymore because of its toxicity. What happens to

cells when too much of the drug is taken? Explain the sequence of events that lead to celldeath.


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5) (11-39) When membrane proteins are purified and reconstituted into artificial

membrane vesicles, they can be inserted in either their normal orientation or inverted.This means that a reconstituted pump is pumping into some vesicles and pumping out of

others depending how in inserted. You have developed a sorting procedure that will

allow you to purify vesicles that only have a Na+/K+ pump oriented so that the part that

normally faces the cytosol is on the outside of the vesicle. Predict what would happenunder the following conditions:

A) Solution inside and outside has Na+ and K+, but no ATP

B) Solution inside the vesicles contains both Na+ and K+, the solution outside has both

ions plus ATP.

C) Solution inside has Na+; solution outside has Na+ and ATP

D) The same solution as in B), but before you purified your oriented vesicles.

6) The gastric parietal cell secretes H+ into the stomach to make stomach acid. If you are

constantly vomiting, secretion is ramped up to refill the stomach with acid. This is theinitiating event that can cause a number of problems for people with bulimia. The

diagram describes the pumps and channels in the gastric parietal cell and their placement.

H+ and HCO3- are generated from H2O and CO2 by carbonic anhydrase.A) Label all the membrane proteins an ion channel, a pump, a symporter, or an antiporter.

B) Is transport by the Cl-/HCO3 transporter energetically favorable or unfavorable? If itis unfavorable, explain where the energy is coming from to power transport.

C) Explain what would happen to intracellular pH if you treated gastric parietal cells witha drug that inhibits each of the following: i) H+/K+ pump

ii) Cl-/HCO3 transporter

iii) carbonic anhydrase

D) Using this diagram, explain what constant vomiting does to blood pH and how thatchange occurs.

E) Using the diagram, does vomiting cause any additional changes in electrolytes in thebloodstream (Na+, K+, Cl-, etc). Explain your reasoning.


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7) GLUT1 is a glucose transporter found in many cells. The Kmof GLUT1 for D-glucoseis 1.5mM, and the Vmax for GLUT1 transport of glucose is 500 !moles transported per

milliliter of cells.

A) Plot velocity of glucose transport vs. extracellular glucose concentration.

B) Is GLUT1 transport of glucose an example of facilitated or passive diffusion across

the plasma membrane? Draw what the other kind of transport would look like on

your graph.

C) How would this graph change if the cells expressed only a partially functional mutant

version of GLUT1 with a higher Km of 7.5mM for glucose?

D) How would this graph change if the cells expressed wildtype GLUT1, but only hadhalf as many transporters on their surfaces?


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E) There is a high free energy barrier to transporting glucose across the plasma

membrane in the absence of GLUT1 and other transporter molecules. Explain the causeof this free energy barrier.

F) Transporters lower the energetic barrier and facilitate transport across the membrane in

both directions. If transporters are not unidirectional, what causes glucose to betransported from outside to inside cells instead of vice-versa?

G) When GLUT1 transporters are reconstituted in liposomes and then incubated withglucose, the levels of glucose transport into and out of the liposomes reaches a state of

equilibrium. Why do erythrocytes in the body that absorb glucose not achieve

equilibrium between import and export of glucose? [Hint: What happens to glucoseonce

it enters these cells?]

8) You have developed a fluorescence based flippase/scramblase assay. This assay

utilizes 1) phospholipids that have been labeled with the fluorescent molecule NBD and2) the ability of dithionite to reduce the NBD labeled phospholipid to non-fluorescentamine products. Flippase/Scramblase activity is measured by the loss of fluorescence as

lipids are transferred to the outer leaflet. The assay is diagrammed below:

You have decided to study a scramblase that is specific

for phosphatidylcholine by making vesicles that have thescramblase, NBD-phosphatidylcholine and NBD-

phosphatidylserine. You make the vesicles and at time

A you add dithionite and at time B you add detergent.

The solid line is the data from vesicles withoutscramblase and the dotted line is the data with the

scramblase.

A) Why does the lipid-only vesicle decrease to half itsfluorescence when dithionite is added?

B) Why does addition of detergent cause the

fluorescence to go to zero?


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C) Explain why the scramblase vesicles end up at a lower fluorescence than pure lipid

vesicles?

D) Explain why the scramblase vesicles dont completely bleach? How could you test

your idea?

9) You have prepared lipid bilayers containingN-palmitoyl ceramide

phosphoethanolamine (PCPE) and compared its membrane behavior with D-erythro-N-

palmitoyl-sphingomyelin (PSM). You have focused on membrane fluidity as measured

by fluorescence anisotropy (Moreanisotropy means less membrane

movement/fluidity). You measure the

membrane fluidity of each bilayer with and

without cholesterol under a range oftemperatures and get the following graphs.

A) For pure PSM and PCPE bilayers, labelwhich parts of the curve the membrane isliquid and which parts is it gel-like.

B) What is the phase transition temperature

for PSM and PCPE?

C) Which lipid (PSM or PCPE) would you

want more of as temperature drops? Explainyour reasoning.

D) When cholesterol is present, thetemperature behavior of both membraneschanges. What changes do you observe and

explain why cholesterol has these effects?

bicd 110 wi15 problem set#1

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