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BL A512 -- Signal Transduction Fall -- 1998 -- Prof. Stark
Midterm Exam -- March 3, 1998 -- 100 points

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1. Name four amino acids which are phosphorylated in signalling systems in eukaryotes and prokaryotes? (4 points)
tyrosine, serine, threonine, histidine, aspartate

2. Show with an equation and / or a graph how the unit Siemens is related to Ohm's law. (2 points)
if E=IR (Ohm's law), thinking of E as the y-axis and I as the x-axis, R is the slope
so for I=gE, g ([1/R] in Siemens) is slope

3. Where in the G-protein linked receptor are the (a) glysosylation (b) G-protein interaction and (c) phosphorylation sites? (3 points)
(a) near N-terminus, outside cell
(b) the second & third cytoplasmic loops and the C-terminus
(c) near C-terminus

4. Draw the general structures of insulin (and how that structure relates to proinsulin) and the insulin receptor (and how its subunits are positioned in the membrane. (3 points)
insulin, see Alberts et al., Figs. 3-14 and 3-54
insulin receptor, see Fig. 15.47

5. For voltage to gate activation of the action potential's sodium channel or the Shaker potassium channel, what happens to what part of the molecule? (2 points)
S4, with one chrged arginine evert 3-4 amino acids (1 per alpha helix period)
rotates and moves

6. What kind of a "filter" (in terms of the frequency pass of the RC circuit) is a glass micropipette and why is this time constant so high in terms of the duration of fast signals like the action potential? (2 points)
it's a low pass filter, very slow because the resistance of such a small tip is
high and the capicitance of glass is also high

7. What molecule donates the phosphate to phosphorylate the sodium potassium pump? (1 point)
ATP

8. Why is the potential calculated by the Nernst equation called an equilibrum potential? (1 points)
equilibrium is assumed, which works out to chemical and electrical potentials
being equal and opposite
9. What is the term used to describe the kind of mechanoreceptor in the fingertips which would be used to detect the roughness of a surface while running the fingertips over that surface? (1 point)
phasic which means it is rapidly adapting

10. Where is dystrophin in relation to a mechanogated channel? How did dystrophin get its name? (2 points)
involved in hooking channel to intracellular actin
from muscular dystrophy, specifically Duchenne or Becker

11. What are the two interesting intracellular domains of the CFTR? (2 points)
nucleotide (ATP) binding folds, R-domain: charged phosphorylation consensus sequences

12. What were the general strategies used in the first isolations of the sodium channel from Electrophorus and the Shaker potassium channel from Drosophila. To find a mouse homologue of the Drosophila Shaker channel, what is done? (3 points)
for Electrophorus, get lots of protein, sequence, probe,
for Shaker, get conditional mutants, for homologue, low stringency hybridization

13. SNARE stands for SNAP receptor. Where (cellular compartments) are SNARE and SNAP? (2 points)
there are v-SNARE (vesicle) and t-SNARE (target=membrane
SNAP (S=soluble) is cytoplasmic

14. Compare the concentration (number per unit of area) of nicotinic acetylcholine receptors at the end plate with the axonal sodium channel. (2 point)
real high (20,000 - 30,000 per square micron) vs real low (like 13 per square micron)

15. What happens to the conductance of the chloride channel during an inhibitory post-synaptic potential (IPSP) triggered by an inhibitory neurotransmitter like GABA? (1 point)
the conductande goes up

16. What does botulism toxin do in the nervous system and what protein is specifically affected? (2 points)
binds synaptobrevin=VAMP=v-SNARE to inhibie vesicle release

17. Here is part of a diagram redrawn from Berridge's 1985 Scientific American paper on intracellular communication. Draw a corrected diagram for receptor and G-protein taking into account present knowledge about membrane localization and subunit structure. (3 points)
the receptor crosses the whole membrane
G-protein is heterotrimeric (alpha, beta and gamma), the alpha and gamma anchored
to the membrane by fatty acids, and thus the protein is just inside the membrane









18. What two molecules are ADP-ribosylated by the actions of what two respective toxins, and what does that do to the action of those two molecules? (4 points)
G alpha s (stimulatory) by cholera blocks GTP hydrolysis, continuous activation
G alpha i (inhibitory) prevents ADP release, cannot be activated


19. Discuss the NMDA receptor with respect to (a) type (e.g. enzyme, G-protein linked, ionotropic, etc), (b) the way this receptor activates neuronal nitric oxide synthase, and (c) the mechanism of facilitating the receptor - NOS interaction. (3 points)
it is a channel, calcium through channel activated NOS via CaM
PSP protein holds them together by PDZ domains

20. The small GTP binding protein Ras requires a GEF (guanine nucleotide exchange factor, alias GNRP = guanine nucleotide releasing protein) and a GAP (GTPase activating protein). By contrast, how does the alpha subunit of the heterotrimeric G protein perform these functions? (2 points)
GEF function performed by activated G-protein coupled receptor
GAP = GTPase activating protein

21. What is unusual about cAMP signalling as the Dictyostellum aggregation pheromone system as compared with the Gs system? In other words, compare location of action and molecule with which cAMP interacts. (4 points)
cAMP is outside vs inside the cell
cAMP activates G-protein coupled receptor vs A-kinase

22. How is a CFTR homologue related to yeast mating? (1 point)
a-factor (pheromone) is secreted by such a multi drug resistance transporter

23. What is the membrane receptor for the subunit of cholera toxin which facilitates the entry into the cell of the other subunit which interacts with the G protein? (1 point)
ganglioside GM1

24. What alteration is made to what amino acids on what portion of the bacterial chemoreceptor protein to mediate adaptation at the receptor level? (3 points)
methylation, glutamate, 4 times intracellularly


25. Describe the geometry of the protein used in gap junctions from the standpoint of number of membrane spans per protein, number of proteins per unitary channel, relationship of such channels in two communicating cells, and arrangement (number or geometry) of unitary channels per gap junction. (4 points)
4 alpha helix membrane spans per molecule, N- and C-termini intracellular
6 molecules per channel, in register with that of adjacent cell
a macular patch of these in the 2 communicating neighbors

26. What enzyme makes an amino acid derivative into a monamine neurotransmitter such as a catecholamine? (1 point)
an aromatic amino acid decarboxylase

27. What molecule mediates the calcium dependence of the potassium channel of Paramecium . By contrast, what is the calcium-binding protein mechanism for vesicle release? (2 points)
calmodulin, synaptotagmin

28. Describe the neurotransmitter ligand channel. For instance, what is the configuration of the membrane spanning region(s) of each protein subunit? For the nicotinic acetylcholine receptor, what is the arrangement of subunits? (3 points)
4 spans per molecule, N- and C-termini outside cell
5 subunits, for nicotinic, 2 alphas, beta gamma, delta

29. How does cAMP regulate PKA? How is cAMP's action ended, and what common drug inhibits this process? (3 points)
2 cAMPs each pull 2 inhibitory subunits off 2 catalytic subunits
phosphodiesterase, caffeine

30. Where is the IP3 receptor located? (1 point)
on the membranes of clcium sequestering intracellular reticula

31. Where is adenylyl cyclase located and what causes it to be located there? (2 points)
it is a membrane protein with 2 portions each with 6 transmembrane alpha helices

32. In bacterial chemotaxis, ligand (aspartate) activates receptor (Tar) which, in turn activates kinase regulator (cheW) to regulate autophosphorlation of cheA. Where does CheA get its phosphate from, and what two molecules does it donate its phosphate to? (3 points)
from ATP, to cheY and to cheB

33. Give two of the original pieces of evidence that the chemoreceptor system in bacteria did not depend on metabolism of nutrients? How many different types of receptors are there now thought to be in E. coli? (3 points)
some metabolized molecules do not attract, non-metabolized molecules can attract
attraction evn if another nutrient is present, etc

34. What would a disorder of the human homologue of the ether-a-go-go gene's potassium channel do to the electrocardiogram? (1 point)
long QT syndrome, QRT is ventricular depolarization, T repolarization -> long act. pot.'s

35. How (if at all) does the G protein differ in signalling pathways which involve PKA vs PKC? (2 points)
Gs vs Gq


36. What is the relative size of the alpha subunit of the heterotrimeric G protein vs. the ras protein? (1 point)
even just the alpha subunit is much bigger than ras

37. Carbon monoxide is synthesized from heme. By contrast, what is the precursor for nitric oxide? Also, by contrast, what does heme have to do with nitric oxide synthesis? (2 points)
from amino acid arginine
it is the active site in the NOS enzyme

38. What would happen to the end plate potential and why if the concentration of extracellular calcium were decreased? (2 points)
lower, even to miniature end plate potentials (1 per vesicle), calcium needed for release

39. Behaviorally, how does a bacterium wind up approaching an attractant? (1 point)
longer runs when going in right direction, more tumbles in wrong

40. What is unusual about the GC involved in the sea urchin spermatozoan attraction to the peptide released by the egg and for atrial natriuretic peptide? (1 point)
it is a membrane receptor

41. In the somatic motor system, the nicotinic receptor is at the end plate. By contrast, in the autonomic motor system, where, if at all, is the nicotinic receptor used? (2 points)
in all (sympathetic and parasympathetic) ganglia

42. What is unusual about how the nervous system mediates regulates the vascular smooth muscle in the corpus cavernosum? What is the mediator derived from the endothelium liberated through the effects of that unusual transmitter mechanism? (2 points)
the parasympathetic nervous system usually does not innervate arteriole smooth muscle
endothelium derived relaxation factor = NO

43. Stock et al. propose that cheY is analogous to ras. What would be the specific analogues of activated ras (ras-GTP), inactivated ras (ras-GDP), and GAP? (3 points)
cheY with phosphate attached, without phosphate, cheZ


44. What are the respective predominant mechanisms for removing acetylcholine vs. norepinephrine from the synaptic cleft? (2 points)
breakdown by acetylcholinesterase, reuptake

45. What is special about arachidonic acid in terms of its structure and its products? (2 points)
long (18), polyunsaturated (4), -> prostaglandins and leukotrienes


46. Give an example of a ligand-gated channel gated from the inside of the cell. (1 point)
rod phototransduction


47. What kind of molecule is a response element part of? (1 point)
DNA in the promoter


48. How much ATP, if any, is utilized to operate the sodium channel which mediates the action potential? (1 point)
none, transport is passive

49. What is muscarine? (1 point)
the agonist to define muscarinic acetylcholine receptors




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